**1. Introduction**

Mycotoxins, secondary fungal metabolites, are frequent contaminants of cereals and cereal products. Delivered via these plants and products, they pose a serious health threat to humans and animals [1–3]. Whereas mycotoxins are sometimes regarded as stressors [4,5], they have a target-specific mode-of-action and so are true toxins rather than stressors per se [6]. The most common and important mycotoxins in Europe, produced by fungi of the *Fusarium* family, are deoxynivalenol (DON), and zearalenone (ZEN). Pigs are particularly sensitive to DON [7], but they also show high sensitivity to ZEN [8].

In pigs, high doses of DON cause reduced appetite, complete anorexia, vomiting [9], and reproductive disorders [10]. Absorption of DON is rapid, and the toxin reaches the peak

plasma concentrations within 30 min of oral administration [11]. The majority of ingested DON is absorbed in the proximal part of the small intestine [12]. In the liver, DON is metabolized into de-epoxy-DON [13]. An evidence showed that chronic ingestion of DON at low doses, which is clinically asymptomatic, alters the mucosal epithelial cells and villi of the small intestine [14–17] and affects the defense mechanisms of the large intestine [18].

ZEN and its metabolites are agonists of estrogen receptors, and they compete with endogenous hormones for the binding sites of estrogen receptors [2,19]. Treatment with ZEN leads to precocious puberty, reproductive disorders, and hyperestrogenism [20]. According to previous studies, domestic pigs are particularly sensitive to the estrogenic effect of ZEN owing to the very rapid and large (approximately 80–85%) absorption of the toxin in their digestive system [21]. In the liver, ZEN is metabolized into α- and β-zearalenol, which are considered more toxic than ZEN. The ratio of these two metabolites to each other is species-specific. Most studies showed that α-zearalenol predominates in pigs [13,21–25]. This form is more active than β-zearalenol, which provides another explanation for the high sensitivity of pigs to ZEN content in feed [21,24]. ZEN at low doses has deleterious effects on the morphology of the small intestine [16,17] and affects the defense mechanisms of the liver [26] and the large intestine [18].

During exposure to environmental mycotoxins, animals often encounter mixtures of toxins, rather than single toxins. Therefore, the toxicity of mycotoxins needs to be addressed in the context of their mixtures to assess their health risk [27–29]. However, studies on the effects of mycotoxin combinations are relatively rare, and their results are very ambiguous [27,28,30,31].

The liver and gastrointestinal parts of the digestive system are major sites of mycotoxin metabolism [32]. However, there is little molecular, metabolic, and histological research on the effects of mycotoxins on the liver. In addition, the outcomes of these studies are not conclusive [15,28]. In pigs, some histological changes have been observed in the liver as a result of the individual or combined effects of DON and ZEN [15,33]. Contrastingly, Renner and coauthors [34] showed that chronic dietary DON exposure (4 weeks, 4.59 μg/kg BW) did not affect the histology of pig liver.

The purpose of this study was to determine the effects of single and combined administrations of DON (12 μg/kg BW) and ZEN (40 μg/kg BW) to pigs for 1, 3, or 6 weeks on the histology and ultrastructure of the liver. The selection of the doses used has been widely discussed in our previously published papers [16–18].

### **2. Results and Discussion**

### *2.1. Light Microscopy Study*

### 2.1.1. Architecture of the Liver

The livers of control pigs had the typical structural characteristic for this species (Figures 1A and 2A). The hepatic lobules were neatly outlined by an envelope of fibrous connective tissue, which interconnected the portal areas. Within each lobule, hepatocytes were arranged in linear cords radiating from the central vein and separated by sinusoids, which had a uniform diameter over their entire length. The limiting plate of hepatocytes bordered the lobule interior from the connective tissue.

Prominent qualitative differences in the liver architecture were observed between the control group and the DON and DON + ZEN groups. The perilobular connective tissue widened and contained greater amounts of collagen fibers starting from the first week of treatment (Figures 1B and 2B, Table 1). In addition, connective tissue penetrated into the lobule and caused disruption of the limiting plate (Figure 1C). After 6 weeks of treatment, the presence of small scars was pronounced (Figure 1D). The disorganization of hepatic cords was observed in the livers of pigs fed diets contaminated with DON and DON + ZEN for 3 and 6 weeks.

**Figure 1.** (**A**,**B**) Architecture of the liver in a control pig, 6th week of experiment (**A**) and in a pig receiving deoxynivalenol (DON) for 6 weeks (**B**). Note the thickening of the interlobular septa. (**C**) A strip of connective tissue in the liver lobule. A pig was treated with DON for 3 weeks. (**D**) Focal fibrosis inside the liver lobule of a pig treated with DON for 6 weeks. Note that the central vein is surrounded by connective tissue. Figures (**A**–**D**) show hematoxylin and eosin stained sections.

**Figure 2.** (**A**,**B**) Trichrome staining of the liver in a control pig, 6th week of experiment (**A**) and in a pig receiving DON for 6 weeks (**B**). Pay attention to the thickness of the interlobular septa and the content of collagen fibers in them.

**Table 1.** Semiquantitative analysis of fibrosis.

Description of the score used: 0, no fibrosis, perilobular tissue septa with small number of collagen, characteristic for the pig liver; 1, mild enlargement of portal areas and perilobular tissue septa; 2, large enlargement of portal areas and perilobular tissue septa, limiting plate fibrosis; 3, partial cirrhosis of some lobules. The values presented are means.

The administration of DON and DON + ZEN significantly increased the thickness of perilobular connective tissue in the liver after 3 and 6 weeks of treatment. No significant differences were observed in the thickness of septa after ZEN treatment (Figure 3A). The cross-sectional area of the lobules did not differ between the control group and the groups treated with mycotoxins (Figure 3B).

The data show that DON and DON + ZEN, but not ZEN, affected the qualitative and quantitative characteristics of the liver architecture. The administration of DON and DON + ZEN resulted in an excessive accumulation of extracellular matrix, including collagen, in perilobular connective tissue, starting from the first week of treatment. Moreover, treatment with DON + ZEN for 3 and 6 weeks caused a progressive accumulation of extracellular matrix in the liver parenchyma. We interpreted the intra-lobular presence of mild scars as early fibrosis. The process of liver fibrosis is most often a consequence of chronic diseases, but it can also be caused by many factors that induce acute damage [35,36]. In experimental studies, administration of xenobiotics induced parenchymal fibrosis similar to cirrhosis in rodent liver [36]. The intensity of the fibrosis process caused by external factors

shows large differences [37]. Previous studies have shown that mycotoxins affect the histology of pig liver [15,34,38]; however, the authors did not report the process of liver fibrosis.

**Figure 3.** (**A**) Thickness of the connective tissue septa. (**B**) Cross-sectional area of the liver lobules. C—control group, Z—group treated with zearalenone (ZEN), D—group treated with deoxynivalenol (DON), and M—group treated with DON and ZEN. The values are presented as mean ± standard deviation. Bars labeled with different small lower-case letters differ significantly at *p* ≤ 0.05.

Disorganization of hepatic cords as one of the main lesions in piglets after chronic exposure to DON was observed by Gerez et al. [15]. In our study, similar changes were found after 6 weeks of DON and DON + ZEN ingestion.

#### 2.1.2. Microscopic Liver Scoring

The analysis using modified microscopic liver scoring (histology activity index (HAI)) comprised six histological criteria. Exemplary microphotographs of changes included in the individual criteria are presented in Figure S1 (Supplementary Material). The cumulative score and the contribution of each criterion are presented in Figure 4. The largest histopathological lesions of the liver were observed in pigs treated with DON for 1 week or DON + ZEN for 1, 3, 6 weeks. The total HAI score for DON resulted mainly from increased portal inflammation and focal lytic necrosis, whereas that for DON +

ZEN resulted from increased portal and periportal inflammation, confluent necrosis, and especially focal lytic necrosis, compared with control pigs. The total HAI scores in pigs receiving DON and DON + ZEN for 1, 3, and 6 weeks were significantly higher than in that in the control group.

**Figure 4.** Histopathological score of the examined livers according to the histology activity index (HAI) modified by Stanek et al. [38]. Six histopathological parameters were scored in hematoxylin and eosin (HE)-stained tissues (see Figure S1 in Supplementary material), and the cumulative HAI score was calculated for each experimental group. The mean value of the cumulative HAI scores in each group is represented by the total height of the bar. C—control group, Z—group treated with zearalenone (ZEN), D—group treated with deoxynivalenol (DON), and M—group treated with DON and ZEN. The values of the cumulative HAI score labeled with different lower-case letters above the bars differ significantly at *p* ≤ 0.05.

The foci of hepatocyte necrosis with lymphocytic infiltrates, portal, periportal, and acinar inflammation occur in the liver as a result of drug usage, intoxication, or viral and bacterial infections [39]. The occurrence of necrosis is also associated with the activation of immune mechanisms in response to the adverse effects of toxic substances [36,40]. These changes are generally associated with pathological symptoms; however, they also occur in seemingly healthy animals, and, in this case, they are considered as preclinical conditions [41].

The results of our HAI analysis showed that the liver, being the first metabolic station, is affected to varying degrees by the administered toxins. Treatment with ZEN had no significant effect on liver histopathology. However, DON and DON + ZEN significantly affected pig liver regardless of the administration period of toxins. The highest intensity of necrosis foci was observed in the livers of animals receiving both mycotoxins. The results obtained may be interpreted as an effect of the toxic effects of DON and the synergistic toxic effect of the combined toxins. In addition, they may be caused by the stimulation of inflammatory processes in response to the administered mycotoxins. This is due to the pro-inflammatory actions of DON and ZEN [42].

The results of previous studies on the effect of mycotoxins on the histology of the liver parenchyma are diverse. Histological changes, including the disorganization of hepatic cords, the cytoplasmic vacuolization of hepatocytes, megalocytosis, and focal necrosis, were reported in pigs subjected to 28 days of diet contaminated with DON (3 mg/kg) or DON (3 mg/kg) + NIV (1,5 mg/kg) + ZEN (1,5 mg/kg) [15]. Acute exposure to DON at a dose of 1 mg/kg BW for 6 and 24 h led to apoptosis of hepatocytes [33]. On the other hand, exposure to DON at a concentration of 4.59 mg/kg feed for 28 days [34] or 3.1 mg/kg feed for 37 days did not affect liver HAI [38].

Megalocytosis and cytoplasmic vacuolization were the main histological lesions reported in piglets subjected to chronic exposure to DON [15]. Magalocytosis potentially indicates irreversible hepatocyte injury [43]. However, in our study, this phenomenon concerned only a few hepatocytes, and there were no differences in their presence between control and experimental pigs.

### 2.1.3. Hepatic Sinusoids

The livers of pigs from all experimental groups were characterized by significant dilatation of the hepatic sinusoids in Zone III (drainage) of acinus compared with those of control animals (Table 2; Supplementary material, Figure S2). The largest sinusoidal dilatation was observed in pigs receiving DON + ZEN. In these animals, significant sinusoidal dilatation occurred after 1, 3, and 6 weeks of mycotoxin administration. In the groups treated with ZEN alone and DON alone, sinusoidal dilatation decreased after 3 and 6 weeks (Table 2).

**Table 2.** Semiquantitative analysis of sinusoidal dilatation.

Description of the score used in estimating changes: 0, absence; 1, sporadic presence; 2, few presence; 3, middle presence; and 4, numerous presence.

Under physiological conditions, the liver sinusoids are characterized by a constant, uniform diameter. Sinusoidal dilatation in Zone I has been observed in pregnancy, long-term steroid administration, and hepatomegaly [44]. This phenomenon also occurs after exposure to vinyl chloride and arsenic [40], and may also be the result of the administration of certain chemotherapy drugs [45]. Sinusoidal dilatation in Zone III was observed in the case of local disorders of blood flow in the portal veins or hepatic veins [40]. In our study, pronounced sinusoidal dilatation in Zone III was observed as a result of DON and ZEN administration, which suggested a disruptive effect of mycotoxins on blood flow in the liver. However, the histological findings did not provide any suggestions about the possible cause of this disturbance.

### 2.1.4. Glycogen Storage

We used periodic acid-Schiff (PAS) staining to evaluate changes in glycogen content. An increase in glycogen deposits was observed in the groups treated with ZEN, DON and DON + ZEN compared with those in the control group after the first week of experiment (Figure 5, Table 3). The staining was particularly intense in Zone I. After 3 and 6 weeks of treatment, there were no differences in glycogen content between the control and experimental groups.

**Table 3.** Semiquantitative analysis of glycogen content in the central and peripheral parts of liver lobules.

For a description of the score, see Table 2.

 The first week of experiment. Periodic acid-Schiff reaction. Note the increase in the amount of glycogen in a pig treated with mycotoxins.

The liver is a major site of glycogen accumulation, and the main role of glycogen in the liver is to store glucose for release during fasting. The amount of stored glycogen depends on nutritional and living factors and shows large individual fluctuations [36,40]. In previous studies, no effects of mycotoxins on glycogen storage were observed in pig liver [7]. The results of our current studies showed that the effect of DON and ZEN on liver glycogen was temporary.


Author details

The mechanism involved in the ovarian diathermy is that it leads to the correction of hypersecretion of LH brought about by modification of the ovarian pituitary feedback. The practice is to drill into both ovaries. However, unilateral drilling has been found to bring about ovulation in either ovaries as well [1, 15, 26]. A net meta-analysis comparing its use as an ovulation induction agent did not find it superior to placebo or no treatment. It had no significantly higher chance at pregnancy or higher pregnancy rates in women with PCOS [6]. In patients with clomiphene resistance where it is being advocated, LOD was found to reduce testosterone and Luteinising hormone levels and associated with more regular cycles, higher ovulation and pregnancy rates compared to metformin alone even though metformin results in more attenuation of luteinizing hormone [28]. Emerging evidence shows that unilateral ovarian drilling has similar effects as bilateral ovarian drilling in bringing about ovulation, pregnancy rates, and life birth rates. Reported pregnancy rates are lower than in treatment with HMG [24]. The seeming comparative advantage of LOD is in its one-off therapy, especially in patients with clomiphene citrate resistance, sustained reversal of the pathology, high ovulation and pregnancy rates, reduced risk of multiple pregnancy, and ovarian hyperstimulation syndrome as well as patient's acceptability [26]. LOD with elecrocautery was found to be superior in treating anovulation compared to laser drilling or use of gonadotropins in clomiphene-resistant PCOS patients. The major side effects of LOD are the fact that it is a theater procedure and requires anesthesia; it may reduce ovarian reserve and has been associated with peri-ovarian adhesions [29, 30]. When ranked according to efficacy of ovulation induction, the systemic review found out that the clomiphene citrate in combination with metformin was the most efficacious followed by follicle-stimulating hormone, letrozole, metformin, clomiphene, tamoxifen, laparoscopic ovarian drilling, and placebo or no treatment in that order. This when ranked in percent-

age efficacy of effectiveness showed 90, 82, 80, 50, 46, 27, 22 and 3%, respectively [6].

Bariatric surgery has been used for weight reduction among highly obese women who had bariatric surgery for just weight reduction. The bariatric surgery in obese PCOS patient also

However, when the ranking in terms of live birth rate was done, letrozole (81%) gave the best result, followed by follicle-stimulating hormone (74%), clomiphene-metformin (71%), tamoxi-

Women with PCOS should undergo pre-conception counseling before any treatment for infertility. The importance of life-style modification, especially weight loss and exercise, should be emphasized and encouraged in overweight women, and smoking and alcohol consumption should be discouraged [24]. More randomized trial to determine the effect of weight loss to ovulation should be undertaken to elucidate clearly the place of weight loss as a means of ovulation of induction considering its affordability and acceptability as a means of treatment.

fen (48%), clomiphene citrate (36%), metformin (30%), placebo or no treatment (10%).

4.2. Use of bariatric surgery to induce ovulation

5. Conclusion

84 Debatable Topics in PCOS Patients

resulted in weight loss, spontaneous ovulation, and pregnancy.

Vaduneme K. Oriji\* and Kennedy Nyengidiki

\*Address all correspondence to<EMAIL_ADDRESS>
University of Port Harcourt and University of Port Harcourt Teaching Hospital, Port Harcourt, Nigeria


**8. Conclusions**

Foodborne diseases are one of the main causes of morbidity and mortality, especially in low-income countries having poor sanitation and inadequate healthcare facilities. Several plant extracts and phytochemicals including catechin, epigallocatechin, apigenin, kempferol, berberine, tiliroside and quercetin were found to be highly e ffective to control the growth of these foodborne pathogens causing infections in in vitro systems. Various mechanisms have been reported for the antimicrobial activities of medicinal plants and they a ffect those pathways of macromolecular metabolism which are the proven targets for antibiotic intervention. Medicinal plants can impact more than a single specific antimicrobial target as they contain a large number of biochemical phytoconstituents. Once utilized, natural products with antimicrobial properties can be e ffective for the prevention and treatment of foodborne diseases and can increase the shelf life of food products. However, most of the studies covered in this review are performed using in vitro assays. Further detailed in vivo studies for exploration of e ffectiveness and mechanism of their activities are necessary. Along with that, most of these studies were performed in the in vitro systems without the addition of food, simulated gastric and/or intestinal juice. Various such factors may alter the activity of these extracts and compounds when used in complex biologic systems such as in vivo and human studies. Future studies should also focus on the pharmacokinetic and toxicological aspects of plant extracts and phytochemicals.

**Author Contributions:** F.U. (Farhat Ullah), F.U. (Farman Ullah), M.A., A.A.-D., A.S., I.H. conceived the idea, performed literature review, data collection, interpretation and manuscript preparation. M.S., Z.Y., A.A.-D. and H.P.D. revised the manuscript. All authors have read and agreed to the published version of the manuscript.

**Funding:** There is no particular funding to support this research.

**Conflicts of Interest:** The authors declare no conflicts of interest.


**5. B cell targeted therapy in systemic vasculitis**

#### **5.1. The role of B cells in vasculitis**

The WGET (The Wegener Granulomatosis Etanercept Trial) was a double-blind study that aimed to assess the role of etanercept in the induction and maintenance of remission in 180 patients with GPA. In this trial the use of etanercept showed no impact on the rate of achieving sustained remission compared to placebo (69% for etanercept vs 75% for placebo) with a slight increase in the rate of solid tumors among patients receiving a combination therapy of cyclophosphamide and etanercept. A risk that proved to be insignificant. [31] Another open label phase II prospective study including 14 patients with acute flares of AASV either as first manifestation of disease or relapse demonstrated that the addition of adalimumab to predni‐ solone and cyclophosphamide for the treatment of severe ANCA associated systemic vasculitis (AASV) was associated with response rates and adverse events similar to standard therapy

Infliximab at a dose of 5mg/kg, showed efficacy in cases with refractory uveo-retinitis with improvement of visual acuity, healing of oro-genital ulcers, healing of intestinal lesions and remission. It has also been disclosed that infliximab is a rapid and effective therapy for sightthreatening panuveitis in Behçet's disease. Infliximab administration thus leads to a rapid and effective suppression of acute ocular inflammation, and the remission of the uveitis remained for as long as 28 days after infliximab administration in all five patients. Etanercept is also now being used in refractory Behçet's disease. [33, 34, 35, 36, 37] Despite that Tumor necrosis factor therapy is not an approved therapeutic alternative in Behcet's syndrome due to lack of randomized controlled studies, yet TNF inhibition might provide a valuable alternative in Behçet's syndrome patients who were proven refractory to more commonly used treatments.

TNF has been suggested as an important cytokine in the active phase of coronary disease in mouse models of Kawasaki disease.However, the current use of anti TNF therapy in Kawasaki

Cutaneous vasculitis, is a disease with an annual incidence rate ranging from 39.6 to 59.8 per million, can be classified as primary or idiopathic; or secondary, when it presents as a mani‐ festation of connective tissue diseases, infections, drug reactions or malignancies. [41, 42] Most of the idiopathic cases are self-limited and responsive to supportive measures (limb elevation, warming, avoid standing) and nonsteroidal anti-inflammatory drugs, potent immunesuppressants are sometimes required for the management of the refractory situations. [42]

The high serum levels of the pro-inflammatory cytokines TNF-*α*, IL-1*β* observed in sera of patients with small vessels cutaneous vasculitis supports a potential role of these cytokines in

remains restricted only to cases with immunoglobulin resistant disease. [38,39]

**4.3. Deep idiopathic small vessels cutaneous vasculitis**

alone but with a reduced prednisolone exposure. [32]

248 Updates in the Diagnosis and Treatment of Vasculitis

**4.1. Behcet's syndrome**

**4.2. Kawasaki disease**

**4. TNF-α inhibitors in other forms of systemic vasculitis**

B lymphocytes are key players in immune mediated vasculitis representing the the humoral arm of the immune response. B cells produce pathogenic autoantibodies and because they have multiple effector functions, including antigen uptake and transport, antigen presentation and costimulation of T cells via membrane associated molecules, production of cytokines and chemokines migration to sites of inflammation. B lymphocytes arise from hematopoietic stem cells in the bone marrow. These cells mature independently of an antigen first into pro-B cells, then into pre-B cells and immature B cells. They subsequently enter the antigen-dependent phase in the peripheral lymphoid tissues, where mature-but-naive B cells, after encountering their antigen in the extrafollicular regions of the lymphoid organs, become activated B cells and migrate to the follicular regions. B lymphocytes then exit the follicular regions to differ‐ entiate into memory B cells, late plasmablasts and plasma cells. Specific markers, such as CD20, CD27, BAFF-R (B-cell-activating factor receptor), CD38 and CD138, identify the transitional phases of B cells from stem cells to plasma cells. (Figure 7) [48, 49, 50]

#### **5.2. B cells surface target molecules in vasculitis**

#### *5.2.1. CD-20 cell surface molecule*

CD20 is a 297-amino acid activated glycosylated trans-membrane phosphoprotein specifically expressed on the surface of B cells, starting at the early pre-B cell stage and persists until the differentiation of B cells into plasma cells. CD-20 is not expressed on hematopoietic stem cells, pro-B cells, or normal plasma cells. Plasma-blasts and stimulated plasma cells may express CD20. CD20 is co-expressed on B cells with CD19, another B cell differentiation marker. CD20 appears to play a crucial role in B cell development, differentiation, proliferation and cell-cycle regulation events. B cell mediated disorders with clonal B cell expansion including lympho‐ mas, leukemias, autoimmune diseases were found to be associated with increasing expression of the CD-20 antigen in variable densities. [51, 52, 53, 54]

#### *5.2.2. CD-22 cell surface molecule*

CD22 is a 135-kDa trans-membrane sialoglycoprotein, a member of the immunoglobulin superfamily. Its expression is restricted to lymphocytes of the B cell lineage and is highly developmentally regulated.

CD22 is present in the cytoplasm of pro- and pre-B cells and becomes detectable on the cell surface only at mature stages of B cell differentiation. Cell surface expression is lost during terminal differentiation into plasma cell and after B cell activation. CD22 is also expressed by the vast majority of B cell NHLs. The CD22 molecule has multiple ligands because it binds to α2–6-linked sialic acid residues present on glycoproteins expressed by activated T and B cells, monocytes, neutrophils, erythrocytes, and activated endothelial cells. Although its function is not yet well understood, CD22 appears to be involved in the regulation of B cell activation through BCR signaling, (demonstrating both positive and negative roles in vitro) as well as in cell adhesion. In vivo, the important biological functions of this receptor have been demon‐ strated by genetic disruption of CD22. CD22-deficient mice have a shorter life span, a reduced number of mature B cells in the bone marrow and in circulation, and a chronic exaggerated antibody response to antigen and develop elevated levels of autoantibodies, suggesting a key role for CD22 in B cell development, survival, and function. [55-60]

**5.3. B-cell targeted therapy**

*5.3.1. The B-cell depletion therapy*

*<IP_ADDRESS>. Rituximab*

54] Figure 9.

the use of B cell depletion therapy in vasculitis.

**Figure 9.** Mechanism of B cell depletion with anti-CD20 therapy [54]

Currently available B cell targeted biologic drugs can be classified into two different mecha‐ nisms, the first mechanism is the B cell depletion therapy, the second mechanism acts via inhibition of B cell maturation. Different trials and case reports showed promising results with

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A chimeric human/mouse IgG1 antibody directed at human CD20, which is found on only pre-B and mature B cells. B-cell depletion with rituximab might be useful for patients with autoimmune diseases driven by autoantibody production. Rituximab (anti-CD-20 therapy) showed efficacy in autoimmune disorders including refractory systemic lupus with nephritis and vasculitis. The types of vasculitis investigated in this regard include cryoglobulinemic vasculitis, ANCA associated vasculitis, cutaneous vasculitis with connective tissue diseases like rheumatoid arthritis, systemic lupus erythematosus and Sjogren's syndrome. [51, 52, 53,

#### **5.3. B-cell targeted therapy**

CD22 is present in the cytoplasm of pro- and pre-B cells and becomes detectable on the cell surface only at mature stages of B cell differentiation. Cell surface expression is lost during terminal differentiation into plasma cell and after B cell activation. CD22 is also expressed by the vast majority of B cell NHLs. The CD22 molecule has multiple ligands because it binds to α2–6-linked sialic acid residues present on glycoproteins expressed by activated T and B cells, monocytes, neutrophils, erythrocytes, and activated endothelial cells. Although its function is not yet well understood, CD22 appears to be involved in the regulation of B cell activation through BCR signaling, (demonstrating both positive and negative roles in vitro) as well as in cell adhesion. In vivo, the important biological functions of this receptor have been demon‐ strated by genetic disruption of CD22. CD22-deficient mice have a shorter life span, a reduced number of mature B cells in the bone marrow and in circulation, and a chronic exaggerated antibody response to antigen and develop elevated levels of autoantibodies, suggesting a key

role for CD22 in B cell development, survival, and function. [55-60]

**Bone marrow Antigen Independent**

250 Updates in the Diagnosis and Treatment of Vasculitis

**Pro‐B cells**

**Serum BAFF and CD38**

**Pre‐B cells**

**Immature B cells**

**Mature B cells**

**Cell Surface Antigens**

**Figure 8.** Surface molecules and receptors involved in B cell Maturation, the black bullet refers to identified therapeu‐

**Short lived Plasma cells**

> **Active B cells**

**Memory B cells**

**Peripheral Lymphoid tissue Antigen Dependent Phase**

> **Late plasma‐ blast cells**

**Plasma cells**

**Early Plasma‐blast CD20 CD27 CD40L**

**Stem cells**

**CD19 CD20 CD27**

**CD138**

tic targets [51]

Currently available B cell targeted biologic drugs can be classified into two different mecha‐ nisms, the first mechanism is the B cell depletion therapy, the second mechanism acts via inhibition of B cell maturation. Different trials and case reports showed promising results with the use of B cell depletion therapy in vasculitis.

#### *5.3.1. The B-cell depletion therapy*

#### *<IP_ADDRESS>. Rituximab*

A chimeric human/mouse IgG1 antibody directed at human CD20, which is found on only pre-B and mature B cells. B-cell depletion with rituximab might be useful for patients with autoimmune diseases driven by autoantibody production. Rituximab (anti-CD-20 therapy) showed efficacy in autoimmune disorders including refractory systemic lupus with nephritis and vasculitis. The types of vasculitis investigated in this regard include cryoglobulinemic vasculitis, ANCA associated vasculitis, cutaneous vasculitis with connective tissue diseases like rheumatoid arthritis, systemic lupus erythematosus and Sjogren's syndrome. [51, 52, 53, 54] Figure 9.

**Figure 9.** Mechanism of B cell depletion with anti-CD20 therapy [54]

#### *<IP_ADDRESS>. Epratuzumab*

Epratuzumab is a humanized monoclonal antibody targeting CD22 receptors on B lympho‐ cytes. Two multicenter, placebo-controlled, randomized, double-blind studies (EMBODY™ 1 and EMBODY™ 2), designed to evaluate the efficacy, safety, tolerability, and immunogenicity of Epratuzumab in patients with moderate to severe SLE, are being designed and processed, each will enroll 780 subjects and will last a maximum of 54 weeks. Ongoing experimental studies addressing the role of anti-CD22 in ANCA vasculitis are being run with promising results, whilst randomized controlled trials are still lacking. [61-65]

genotype 1, and interferon induced many side effects. On the other hand, treatments such as cyclophosphamide and plasmapheresis are generally reserved for life- or organ-threatening

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Rituximab showed efficacy in cases of HCV-mixed cryoglobulinemia. The rationale behind anti-B-cell therapy in mixed cryoglobulinemic vasculitis includes the presence immune complex deposition as the cause of symptoms in hepatitis C virus extra-hepatic syndrome, in which chronic stimulation by hepatitis C virus induces the production of cryoglobulins by infected B cells. [75, 76] Rituximab was successfully used in combination with antiviral agents as well as mono-therapy in HCV cryoglobulinemic vasculitis. Rituximab combined with Peg-IFN-α/ribavirin delete both virus-dependent and -independent B-cell clones. Antiviral therapy alone decreased the memory B cells; whereas in association with rituximab, naive B cells are the main depleted population. The observable delay in B-cell reconstitution after rituximab plus Peg-IFN-α/ribavirin stresses the synergistic action of rituximab and antiviral therapy at the immunologic level. Rituximab shortens the therapeutic interval required for achieving a complete clinical response. Clonal expansion of marginal zone–like IgM+ CD27+ B cells (VH1-69 clonal B) has been recently observed in certain HCV-MC patients. Rituximab with Peg-IFN and ribavirin exerts a synergistic effect on polyclonal B lymphocyte expansion. Rituximab plus Peg-IFN-α/ribavirin was more efficient to suppress both memory and VH1-69 clonal B cells compared with Peg-IFN-α/ribavirin alone. A standard therapeutic dose of 375 mg/m2 weekly for 4 weeks is effective, well tolerated and induces a significant and rapid improvement of clinical signs (purpura, arthralgia, peripheral neuropathy) with a decline of cryocrit in most patients with mixed cryoglobulinemia even in cases resistant to IFN therapy. Rituximab trials emphasized the benefit of the drug in inducing remission in cutaneous vasculitis, cryoglobulinemic glomerulonephritis, cryoglobulinemic neuropathy and in

Complete clinical remission was associated with a significant reduction of RF activity and anti-HCV antibody titers. Relapse might occur in up to 36.1%.Complete immunologic response was higher with the combination of rituximab plus Peg-IFN-α/ribavirin. Rituximab treatment of a renal-transplant patient with de novo HCV-related type III cryoglobulinemic MPGN resulted in clearance of cryoglobulinemia, a decrease in proteinuria without a change in serum creatinine or HCV RNA. [69, 77, 78, 79] Rituximab can cause serum sickness, serum sickness like disease, neutropenia and increased risk of infections, pneumopathy, varicella zoster infection, erysipelas, thrombosis of the retinal artery and cold agglutinin disease, occasional flare of vasculitis. Factors that were found to be associated with increased risk of side effects include high complement activation, higher rituximab doses and elevated levels of cryoglo‐ bulins. An increase of viremia might be observed in responders with rituximab monotherapy with insignificant variation of transaminases or deterioration of liver disease. [80, 81] Table 1

Standard treatment of ANCA-associated vasculitic syndromes (AAVS) is composed of remission induction regimen that involves the use of cyclophosphamide and high-dose glucocorticoids, followed by a remission maintenance regimen using methotrexate, azathio‐

underlying malignant lymphoproliferative disorder. [74, 76, 77, 78]

*<IP_ADDRESS>. ANCA-Associated Systemic Vasculitis (AASV)*

disease.

#### *5.3.2. The role of B cells targeted therapy in systemic vasculitis*

#### *<IP_ADDRESS>. Cryoglobulinemic vasculitis*

Mixed cryoglobulinemia (MC) is a chronic immune complex (IC) mediated systemic small vessel vasculitis, characterized by immune complex deposits (cryo-deposits) and frequent visceral involvement. A frequent synonym of this disease is "cryoglobulinemic vasculitis". [66] The term cryoglobulinemia refers to the presence of a single or monoclonal immunoglobulin Ig or more polyclonal immunoglobulins. Cryoglobulins are classified on the basis of their Ig components. These cryoglobulins precipitate at temperatures below 37°C giving rise to high molecular weight aggregates and re-dissolve on rewarming. Cryoglobulins are found in small quantities in normal serum and are present in variable concentrations in many pathological conditions, including myeloproliferative disorders, autoimmune disorders and several infectious diseases. They are classified into: Type I cryoglobulins (10-15% of cryoglobulins) is comprised simply of monoclonal immunoglobulins, typically IgM but less frequently IgG, IgA, or serum light chains. Individuals with Type I cryos typically have a paraproteinemia (e.g., myeloma, Waldenstrom's macroglobulinemia). Type II cryoglobulins (50-60% of cryoglobu‐ lins) occurs when a monoclonal Ig M recognizes and binds to polyclonal IgG's, accordingly, type II cryos are typically IgM-IgG complexes.Type III cryoglobulins (30-40% of cryoglobulins) are composed of polyclonal Ig M that binds to polyclonal IgG. Type II & III cryoglobulinemia are referred to as "Mixed Cryoglobulinemia", these two types are most commonly associated with hepatitis C virus infection. [67]. The term essential cryoglobulinemia was used to describe cryoglobulinemia without identifiable underlying disease, currently it is clear that most of the patients with essential mixed cryoglobulinemia are chronically infected with HCV [68] The classic pathology in cryoglobulinemic vasculitis is leukocytoclastic vasculitis. The disease presents by Meltzer triad which is a triad of purpura, weakness, arthralgia and/or arthritis in 25-30% of cases, thereafter, a series of multisystem pathologies follow involving peripheral nervous system and the kidneys. Widespread vasculitis involving medium-small sized arteries, capillaries and venules with multiple organ involvement may develop in a small proportion of patients [69-72].

The incidence of hepatitis C infection in mixed cryoglobulinemia ranges from 40 to 90%. [69, 73, 74] The hepatitis C virus infects B cells, resulting in clonal expansion and stimulation of autoantibody production. Eradication of hepatitis C with interferon and ribavirin has been tried, but is often ineffective for controlling extra-hepatic disease,[ 45 ] particularly with genotype 1, and interferon induced many side effects. On the other hand, treatments such as cyclophosphamide and plasmapheresis are generally reserved for life- or organ-threatening disease.

*<IP_ADDRESS>. Epratuzumab*

252 Updates in the Diagnosis and Treatment of Vasculitis

Epratuzumab is a humanized monoclonal antibody targeting CD22 receptors on B lympho‐ cytes. Two multicenter, placebo-controlled, randomized, double-blind studies (EMBODY™ 1 and EMBODY™ 2), designed to evaluate the efficacy, safety, tolerability, and immunogenicity of Epratuzumab in patients with moderate to severe SLE, are being designed and processed, each will enroll 780 subjects and will last a maximum of 54 weeks. Ongoing experimental studies addressing the role of anti-CD22 in ANCA vasculitis are being run with promising

Mixed cryoglobulinemia (MC) is a chronic immune complex (IC) mediated systemic small vessel vasculitis, characterized by immune complex deposits (cryo-deposits) and frequent visceral involvement. A frequent synonym of this disease is "cryoglobulinemic vasculitis". [66] The term cryoglobulinemia refers to the presence of a single or monoclonal immunoglobulin Ig or more polyclonal immunoglobulins. Cryoglobulins are classified on the basis of their Ig components. These cryoglobulins precipitate at temperatures below 37°C giving rise to high molecular weight aggregates and re-dissolve on rewarming. Cryoglobulins are found in small quantities in normal serum and are present in variable concentrations in many pathological conditions, including myeloproliferative disorders, autoimmune disorders and several infectious diseases. They are classified into: Type I cryoglobulins (10-15% of cryoglobulins) is comprised simply of monoclonal immunoglobulins, typically IgM but less frequently IgG, IgA, or serum light chains. Individuals with Type I cryos typically have a paraproteinemia (e.g., myeloma, Waldenstrom's macroglobulinemia). Type II cryoglobulins (50-60% of cryoglobu‐ lins) occurs when a monoclonal Ig M recognizes and binds to polyclonal IgG's, accordingly, type II cryos are typically IgM-IgG complexes.Type III cryoglobulins (30-40% of cryoglobulins) are composed of polyclonal Ig M that binds to polyclonal IgG. Type II & III cryoglobulinemia are referred to as "Mixed Cryoglobulinemia", these two types are most commonly associated with hepatitis C virus infection. [67]. The term essential cryoglobulinemia was used to describe cryoglobulinemia without identifiable underlying disease, currently it is clear that most of the patients with essential mixed cryoglobulinemia are chronically infected with HCV [68] The classic pathology in cryoglobulinemic vasculitis is leukocytoclastic vasculitis. The disease presents by Meltzer triad which is a triad of purpura, weakness, arthralgia and/or arthritis in 25-30% of cases, thereafter, a series of multisystem pathologies follow involving peripheral nervous system and the kidneys. Widespread vasculitis involving medium-small sized arteries, capillaries and venules with multiple organ involvement may develop in a small

The incidence of hepatitis C infection in mixed cryoglobulinemia ranges from 40 to 90%. [69, 73, 74] The hepatitis C virus infects B cells, resulting in clonal expansion and stimulation of autoantibody production. Eradication of hepatitis C with interferon and ribavirin has been

particularly with

tried, but is often ineffective for controlling extra-hepatic disease,[ 45 ]

results, whilst randomized controlled trials are still lacking. [61-65]

*5.3.2. The role of B cells targeted therapy in systemic vasculitis*

*<IP_ADDRESS>. Cryoglobulinemic vasculitis*

proportion of patients [69-72].

Rituximab showed efficacy in cases of HCV-mixed cryoglobulinemia. The rationale behind anti-B-cell therapy in mixed cryoglobulinemic vasculitis includes the presence immune complex deposition as the cause of symptoms in hepatitis C virus extra-hepatic syndrome, in which chronic stimulation by hepatitis C virus induces the production of cryoglobulins by infected B cells. [75, 76] Rituximab was successfully used in combination with antiviral agents as well as mono-therapy in HCV cryoglobulinemic vasculitis. Rituximab combined with Peg-IFN-α/ribavirin delete both virus-dependent and -independent B-cell clones. Antiviral therapy alone decreased the memory B cells; whereas in association with rituximab, naive B cells are the main depleted population. The observable delay in B-cell reconstitution after rituximab plus Peg-IFN-α/ribavirin stresses the synergistic action of rituximab and antiviral therapy at the immunologic level. Rituximab shortens the therapeutic interval required for achieving a complete clinical response. Clonal expansion of marginal zone–like IgM+ CD27+ B cells (VH1-69 clonal B) has been recently observed in certain HCV-MC patients. Rituximab with Peg-IFN and ribavirin exerts a synergistic effect on polyclonal B lymphocyte expansion. Rituximab plus Peg-IFN-α/ribavirin was more efficient to suppress both memory and VH1-69 clonal B cells compared with Peg-IFN-α/ribavirin alone. A standard therapeutic dose of 375 mg/m2 weekly for 4 weeks is effective, well tolerated and induces a significant and rapid improvement of clinical signs (purpura, arthralgia, peripheral neuropathy) with a decline of cryocrit in most patients with mixed cryoglobulinemia even in cases resistant to IFN therapy. Rituximab trials emphasized the benefit of the drug in inducing remission in cutaneous vasculitis, cryoglobulinemic glomerulonephritis, cryoglobulinemic neuropathy and in underlying malignant lymphoproliferative disorder. [74, 76, 77, 78]

Complete clinical remission was associated with a significant reduction of RF activity and anti-HCV antibody titers. Relapse might occur in up to 36.1%.Complete immunologic response was higher with the combination of rituximab plus Peg-IFN-α/ribavirin. Rituximab treatment of a renal-transplant patient with de novo HCV-related type III cryoglobulinemic MPGN resulted in clearance of cryoglobulinemia, a decrease in proteinuria without a change in serum creatinine or HCV RNA. [69, 77, 78, 79] Rituximab can cause serum sickness, serum sickness like disease, neutropenia and increased risk of infections, pneumopathy, varicella zoster infection, erysipelas, thrombosis of the retinal artery and cold agglutinin disease, occasional flare of vasculitis. Factors that were found to be associated with increased risk of side effects include high complement activation, higher rituximab doses and elevated levels of cryoglo‐ bulins. An increase of viremia might be observed in responders with rituximab monotherapy with insignificant variation of transaminases or deterioration of liver disease. [80, 81] Table 1

#### *<IP_ADDRESS>. ANCA-Associated Systemic Vasculitis (AASV)*

Standard treatment of ANCA-associated vasculitic syndromes (AAVS) is composed of remission induction regimen that involves the use of cyclophosphamide and high-dose glucocorticoids, followed by a remission maintenance regimen using methotrexate, azathio‐

studies demonstrated successful treatment of the ANCA-associated vasculitides with rituxi‐ mab. [85-88] These early successes led to two landmark studies that support the use of B-cell therapies for these diseases. The efficacy of rituximab in ANCA associated vasculitis was further studied in 2010 in two randomized controlled trials. Both trials examined the use of

A multicentered randomized, double-blind, non-inferiority study of patients with severe GPA and microscopic polyangiitis. In this trial, 197 patients with either new or relapsing disease were randomized in a 1:1 ratio to receive remission-induction therapy with either oral

received the same glucocorticoid regimen (i.e., up to 3 pulses of 1 g of intravenous methyl‐ prednisolone, and then prednisone 1 mg/kg/day, followed by a protocolized taper). Approx‐ imately 50% of subjects enrolled in this trial had significant renal disease and 28% had alveolar hemorrhage at trial entry, although patients were not eligible for this trial if their serum creatinine exceeded 4 mg/dl or if the patient required mechanical ventilation during the study period. The primary end point of this study was achievement of remission at 6 months in the

The RAVE trial showed that rituximab was non-inferior to cyclophosphamide in inducing remission especially in cases with alveolar haemorrhage and/or glomerulonephritis. Rituxi‐ mab was proven superior to cyclophosphamide in the treatment of relapsing disease in AASV. Rituximab therapy was associated with sustained remission over a period of 6 months with encouragingsteroidsparingeffect.Ingeneral,the frequencyof adverse eventswas the same and correlated significantly with the early use of high doses of glucocorticoids in both groups.

*<IP_ADDRESS>.2. The Randomized Trial of Rituximab versus Cyclophosphamide in ANCA-Associated*

A randomized open-label study that looked at the effectiveness of rituximab for the treatment of 44 patients with newly diagnosed ANCA-associated glomerulonephritis. Subjects were randomized in a 3:1 ratio, stratified by age, diagnosis and baseline renal function. The rituximab group (33 patients) received standard dosing of rituximab (375 mg/m2 weekly for 4 doses) as well as two intravenous cyclophosphamide pulses (15 mg/kg) with the first and third rituximab doses. A third dose of cyclophosphamide was permitted if remission had not been achieved after 6 months of therapy. No maintenance therapy was given in this group. The control group (11 patients) received intravenous cyclophosphamide monthly for 3–6 months followed by azathioprine for remission maintenance. Some subjects also underwent plasma‐ pheresis. Remission was defined as the absence of disease activity for 2 months and relapse was defined as any disease activity after remission had been attained. In this trial, 76% of patients in the rituximab group had a sustained remission at 12 months as opposed to 82% in the cyclophosphamide group. Severe adverse effects were noted 42% of the rituximab patients and 36% of patients in the control group. A total of eight patients died, six of whom had been

weekly for 4 weeks). Both groups

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255

rituximab for patients with ANCA-associated vasculitis.

cyclophosphamide (2 mg/kg/day) or rituximab (375 mg/m2

absence of glucocorticoids.

*Vacsulitis (RITUXVAS) Trial*

*<IP_ADDRESS>.1. The Rituximab for ANCA-associated Vasculitis (RAVE) trial*

**Table 1.** Rituximab treatment in Patients with HCV cryoglobulinemic vascultis.CNI = Calcineurin inhibitor; HCV = Hepatitis C virus; HSV = Herpes simplex virus; MMF = Mycophenolate mofetil; NR = Not reported; S = Steroids.↑: Increase; ↓: Decrease; =: No change. [69]

prine or other antimetabolite therapy. Patients who develop undetectable ANCA titers after treatment are less likely to experience disease relapse than patients who remain ANCA positive. Conventional immunosuppression led to a dramatic improvement in the prognosis of patients with increasing remission reaching up to 70%. Conventional immunosuppression might fail to achieve remission in a substantial minority of patients (25-30%). [82] With the tumor necrosis factor inhibition therapy clearly proven to be non-superior to conventional lines of therapy alternative biologic targets are being extensively investigated in AAVS. [31]

B-cell activation is believed to play an important role in the pathogenesis of the ANCAassociated vasculitis concerning antigen presentation, activation of T cell differentiation to Th1 and Th17 cell types and activating TNF-primed neutrophils, leading to premature degranu‐ lation and resultant endothelial damage. [83] Given the role of B cells in the pathogenesis of this disease, B cell depletion and interruption of B cell dependent T cell regulator function with rituximab represents a potentially attractive treatment alternative. [84] Open label and cohort studies demonstrated successful treatment of the ANCA-associated vasculitides with rituxi‐ mab. [85-88] These early successes led to two landmark studies that support the use of B-cell therapies for these diseases. The efficacy of rituximab in ANCA associated vasculitis was further studied in 2010 in two randomized controlled trials. Both trials examined the use of rituximab for patients with ANCA-associated vasculitis.

#### *<IP_ADDRESS>.1. The Rituximab for ANCA-associated Vasculitis (RAVE) trial*

A multicentered randomized, double-blind, non-inferiority study of patients with severe GPA and microscopic polyangiitis. In this trial, 197 patients with either new or relapsing disease were randomized in a 1:1 ratio to receive remission-induction therapy with either oral cyclophosphamide (2 mg/kg/day) or rituximab (375 mg/m2 weekly for 4 weeks). Both groups received the same glucocorticoid regimen (i.e., up to 3 pulses of 1 g of intravenous methyl‐ prednisolone, and then prednisone 1 mg/kg/day, followed by a protocolized taper). Approx‐ imately 50% of subjects enrolled in this trial had significant renal disease and 28% had alveolar hemorrhage at trial entry, although patients were not eligible for this trial if their serum creatinine exceeded 4 mg/dl or if the patient required mechanical ventilation during the study period. The primary end point of this study was achievement of remission at 6 months in the absence of glucocorticoids.

The RAVE trial showed that rituximab was non-inferior to cyclophosphamide in inducing remission especially in cases with alveolar haemorrhage and/or glomerulonephritis. Rituxi‐ mab was proven superior to cyclophosphamide in the treatment of relapsing disease in AASV. Rituximab therapy was associated with sustained remission over a period of 6 months with encouragingsteroidsparingeffect.Ingeneral,the frequencyof adverse eventswas the same and correlated significantly with the early use of high doses of glucocorticoids in both groups.

#### *<IP_ADDRESS>.2. The Randomized Trial of Rituximab versus Cyclophosphamide in ANCA-Associated Vacsulitis (RITUXVAS) Trial*

prine or other antimetabolite therapy. Patients who develop undetectable ANCA titers after treatment are less likely to experience disease relapse than patients who remain ANCA positive. Conventional immunosuppression led to a dramatic improvement in the prognosis of patients with increasing remission reaching up to 70%. Conventional immunosuppression might fail to achieve remission in a substantial minority of patients (25-30%). [82] With the tumor necrosis factor inhibition therapy clearly proven to be non-superior to conventional lines of therapy alternative biologic targets are being extensively investigated in AAVS. [31] B-cell activation is believed to play an important role in the pathogenesis of the ANCAassociated vasculitis concerning antigen presentation, activation of T cell differentiation to Th1 and Th17 cell types and activating TNF-primed neutrophils, leading to premature degranu‐ lation and resultant endothelial damage. [83] Given the role of B cells in the pathogenesis of this disease, B cell depletion and interruption of B cell dependent T cell regulator function with rituximab represents a potentially attractive treatment alternative. [84] Open label and cohort

**Study**

Sansonno *et al. 2003*

Zaja *et al.* 2003

Roccatello *et al.* ,2004

Quartuccio *et al.,* 2006

Basse *et al.* 2005

Visentini *et al.* 2007

**Patients (number with nephritis)**

254 Updates in the Diagnosis and Treatment of Vasculitis

20 (1)

15 (2)

6 (5)

5 (5)

7 (7) (postkidney transplant)

6 (2)

Increase; ↓: Decrease; =: No change. [69]

**Rituximab dose**

375 mg/m2 weekly × 4 weeks

375 mg/m2 weekly × 4 weeks

375 mg/m2 weekly × 4 weeks; 375 mg/m2 monthly × 2 months

375 mg/m2 weekly × 4 weeks

375 mg/m2 weekly × 2–4 weeks

250 mg/m2 weekly × 2 weeks

**Other treatments**

S (low doses)

S (<0.5 mg/ kg/day)

S (one case)

CNI, MMF and S

**Remission overall (nephritis)**

> 16/20 (1/1)

> 13/14 (1/2)

> > 5/5

5/5 (5/5)

7/7

S 4/6 (1/2) 4/5 (2/2)

**Table 1.** Rituximab treatment in Patients with HCV cryoglobulinemic vascultis.CNI = Calcineurin inhibitor; HCV = Hepatitis C virus; HSV = Herpes simplex virus; MMF = Mycophenolate mofetil; NR = Not reported; S = Steroids.↑:

**Remission Purpura Neuropathy**

> 12/14 (6/12)

12/12 (5/5)

4/4 (5/6)

4/4 (1/2) **Side effects HCV viral load**

↑ responders = nonresponders

> ↑ 2/8 ↑ 1/8 = 5/8

4 unchanged

NR

↓ 2/5 = 3/5

NR NR

Septic fever (1)

Retinal thrombosis (1)

Transient bradycardia (2)

Transient neutropenia (1)

Lethal infection (2, fungal and HSV)

Lethal intestinal infarction

**Relapse (number of cases)**

> 4/16 (>7 months)

6 (3–6 months)

2 (>12 months)

3 (>5, >7 and >12 months)

NR

A randomized open-label study that looked at the effectiveness of rituximab for the treatment of 44 patients with newly diagnosed ANCA-associated glomerulonephritis. Subjects were randomized in a 3:1 ratio, stratified by age, diagnosis and baseline renal function. The rituximab group (33 patients) received standard dosing of rituximab (375 mg/m2 weekly for 4 doses) as well as two intravenous cyclophosphamide pulses (15 mg/kg) with the first and third rituximab doses. A third dose of cyclophosphamide was permitted if remission had not been achieved after 6 months of therapy. No maintenance therapy was given in this group. The control group (11 patients) received intravenous cyclophosphamide monthly for 3–6 months followed by azathioprine for remission maintenance. Some subjects also underwent plasma‐ pheresis. Remission was defined as the absence of disease activity for 2 months and relapse was defined as any disease activity after remission had been attained. In this trial, 76% of patients in the rituximab group had a sustained remission at 12 months as opposed to 82% in the cyclophosphamide group. Severe adverse effects were noted 42% of the rituximab patients and 36% of patients in the control group. A total of eight patients died, six of whom had been randomized to receive rituximab therapy. Adverse events were not lower in the rituximab group as had been expected. [90]

Abbrevaitions: Antineutrophil cytoplasmic autoantibodies; RAVE: Rituximab for ANCA-associated vasculitis; RITUXVAS: Rituximab versus cyclophosphamide in ANCA-associated vasculitis. [55,56]

> of B cells to BCR activation. TACI and BCMA signal through the classic NF-κB pathway and through the Mek (mitogen-activated protein extracellular signal-related kinase) pathway to up-regulate anti-apoptotic and down-regulate pro-apoptotic pathways, and through JNK/p38 (c-Jun N-terminal kinase) to drive class-switching. Survival and reactiva‐ tion of B cell memory is BAFF-independent. Plasma cells express TACI and/or BCMA and their survival can be supported by either BAFF or APRIL. In contrast, B1 cells do not require BAFF or APRIL for survival. BAFF plays an important role in humoral immuni‐ ty. T cell-independent type II responses require the interaction of BAFF 60-mer or membrane BAFF with TAC. This interaction is vital for T cell-dependent immunoglobu‐ lin (Ig)M responses. BAFF is also an essential component of the innate immune response and is induced in myeloid DC by type I interferons (IFNs). BAFF up-regulates Toll-like receptor (TLR) expression, promotes B cell survival and, together with IL-6, promotes Ig class-switching and plasma cell differentiation. Soluble BAFF and APRIL are expressed at high levels in the serum and in the target organs of individuals with established anti‐ body dependent autoimmune diseases. Therapeutic antagonism of BAFF and its homo‐ logue APRIL (a proliferation-inducing ligand) targets an important homeostatic signal for B cell survival and selection (Figure 10). Belimumab and atacicept are two potential therapeutic anatgonists to the BAFF-APRIL pathway for B cell activation that are current‐

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257

**Figure 10.** B cells maturation and antibody production in autoimmune diseases.

ly being investigated. [92-105]

**Table 2.** Demographics of patients enrolled in RAVE and RITUXVAS.

Rituximab may be an effective alternative treatment in newly diagnosed as well as refractory ANCA-associated vasculitis. Wegener's Granulomatosis (granulomatosis with polyangiitis) patients with retro-orbital granulomas tend to be less responsive to rituximab therapy. [91] The great limitation of rituximab is some cases with GPA or microscopic polyangiitis may need to be retreated following initial treatment. New anti-CD20 agents, or agents that attack different B-cell precursors, may overcome this hurdle, and may enable even longer periods of remission.


**5.2 How produce native AMF bio-fertilizer unit?**

Production of native AMF bio-fertilizers unit require appropriate funding with adoption of a good financing strategy, based on various technical-economic parameters including description of the income elements and those of the expenses (**Table 3**). Indeed, the elements of income include the sale of bio-fertilizers, remunerations, publicity and assurance. **Table 4** describes all the expenses and revenues provided by the bio-fertilizer production unit.

## **5.3 Economical challenge of native AMF bio-fertilizer production?**

Production of native AMF production is an agro-industrial investment for each arid region, in some case the product may be exported to similar edapho-climatic areas. Designed production and marketing chain must support each unit in order to guarantee the success of the project. In **Table 4** we have tried to establish an approximate economic study which will allow the investors to have an idea on the economic situation of the project, but these figures may vary from one country

#### **Table 3.**

*Realization coast of native AMF bio-fertilizers unit.*

#### **Table 4.**

*Expenses and revenues of native AMF bio-fertilizers unit.*

to another. The main thing is to ensuring the success of the plant development, potentiates the in situ conservation of the AMF community and preservation of ecosystem stability and biodiversity (**Figure 3**).

**91**

*Native Arbuscular Mycorrhizal Fungi and Agro-Industries in Arid Lands…*

In absence of information about the stapes and strategies of native AMF biofertilizer unit, these information given in **Tables 3** and **4** can help in understanding and progressing of this strategy and its application in degraded arid lands for

*Representative plan of AMF production unit on H form (Adapted from Alamri [4] description).*

Arid land has specific ecological characteristics that confer them special management strategies. As same the different soil throughout the world, soil of arid land present a diversity of AMF that may be exploited to enhance agriculture in these areas. For that in the present chapter we have exposed the most important technic of AMF bio-fertilizer production based on native AMF strains. Founded on what has been explained, recommending the use of conventional pot culture technic present the adequate method adapted for arid lands. We have also given an approximate economical evaluation for coast of building and installing native AMF production unit in arid lands. The conventional method includes optimization of scale of production while using the adequate trap plant, law-coast and environmentally safe. With this technology, the preservation of environment from chemical

*DOI: http://dx.doi.org/10.5772/intechopen.94084*

development of sustainable agriculture.

**6. Conclusion**

**Figure 3.**

*Native Arbuscular Mycorrhizal Fungi and Agro-Industries in Arid Lands… DOI: http://dx.doi.org/10.5772/intechopen.94084*

#### **Figure 3.**

*Mycorrhizal Fungi - Utilization in Agriculture and Forestry*

**Space Surface (m2**

Reception 35.61 91,873,800 Conferences room 31.91 82,327,800 Manager's office 17.3 44,634,000 Engineers office 56.02 144,531,600 Storage room 55.13 13,235,400 Isolation room 40.55 104,619,000 Multiplication room 61.88 524,436,600 Conditioning room 64.6 1,666,668,000 Quality Control room 97.99 252,814,200 AMF greenhouse multiplication 203.27 655,545,750 Sales office 41.63 107,405,400 Store checkout 44.34 114,397,200 Changing room and toilets 73.23 188,933,400 Total 823,46 3,991,422,150

**90**

**Table 4.**

**Table 3.**

Supplementary charge

to another. The main thing is to ensuring the success of the plant development, potentiates the in situ conservation of the AMF community and preservation of

**Expenses (\$) Incomes**

Materials 100,000 Tools 100,000 Transport 100,000

Packaging 9,030,000

Publicity 10%

Salaries Varied between 3,225,000 and

Insurance 10% of the operating budget

bio-fertilizer Construction **4,000,000,000**

40,000

13,500,000

10,320,000/employee

10%

2–3%

12900 millions of dollars /year

in average 24\$/kg of

**) Realization coast (\$)**

Native AMF

Investment costs Construction field 65,000/m2

Technical installation

production

Repair and maintenance

dollars of investment

Production charge Raw material

*Realization coast of native AMF bio-fertilizers unit.*

Total 850 milliards of

ecosystem stability and biodiversity (**Figure 3**).

*Expenses and revenues of native AMF bio-fertilizers unit.*

*Representative plan of AMF production unit on H form (Adapted from Alamri [4] description).*

In absence of information about the stapes and strategies of native AMF biofertilizer unit, these information given in **Tables 3** and **4** can help in understanding and progressing of this strategy and its application in degraded arid lands for development of sustainable agriculture.

#### **6. Conclusion**

Arid land has specific ecological characteristics that confer them special management strategies. As same the different soil throughout the world, soil of arid land present a diversity of AMF that may be exploited to enhance agriculture in these areas. For that in the present chapter we have exposed the most important technic of AMF bio-fertilizer production based on native AMF strains. Founded on what has been explained, recommending the use of conventional pot culture technic present the adequate method adapted for arid lands. We have also given an approximate economical evaluation for coast of building and installing native AMF production unit in arid lands. The conventional method includes optimization of scale of production while using the adequate trap plant, law-coast and environmentally safe. With this technology, the preservation of environment from chemical

fertilizer pollution must be enhanced, which lead to economic development of agroindustry adapted for each country with operative system for sustainable agriculture.


**5. Discussion**

This study set out to examine the role of various goals and motivational beliefs on learning among distance learners. It adds to the distance learning literature by examining different goals that distance learners hold and the complex interaction relationship between motivational beliefs and goals.

As expected, both efficacy and control beliefs were significant in predicting the use of learning and regulatory strategies, except for time management strategies. These results indicate that for this group of distance learners a strong sense of efficacy and control beliefs was associated with an engagement pattern that was characterised by adaptive strategy use. These two motivational beliefs also predicted distance learners' attitudes towards learning indicating that a stronger sense of efficacy and control beliefs was associated with learning enjoyment, interest and valuing of learning. To be motivated, distance learners need to feel efficacious and certain about their learning progress.

Another major aim of this study is to examine the moderation effects of efficacy and control beliefs on the impact of goals on strategy use and attitudes towards learning. This study found nonsignificant interaction effect between self-efficacy and various goals included in this investigation. In other words, a strong sense of efficacy beliefs would not enhance the positive effects of adaptive goals such as mastery and performance-approach goals. Neither would a strong sense of efficacy beliefs dampen the negative effects of less adaptive goals such as extrinsic work goals and social enhancement goals.

It should be pointed out that contrary to Dweck's experimental study (Dweck, 1986; Elliot & Dweck, 1988), the current study examined distance learners' perceptions of learning at a rather general level. Dweck's study found significant interaction between self-efficacy and performance-approach goals on learning by requiring students in an experimental setting to complete clearly-defined academic tasks. Subsequent studies (e.g. Harackiewicz *et al.*, 2000; Kaplan & Midgley, 1997; Miller et al., 1993) using correlational design examining the moderation hypotheses at a rather general level did not found significant interaction between self-efficacy and performance goals on learning and achievement. The current nonsignificant results aligned with these correlational studies.

While the level of task specificity may be taken as an explanation for contradictory results between Dweck's study and the current study, a more critical factor may have been the

negatively the use of effort management strategies (=-.20, *p<*.001 controlling for both efficacy and control beliefs). These results suggest that these social goals will lead distance learners to reduce their effort in learning by using surface strategies. Unexpectedly, these

Taken together, regression analyses found that mastery-development goals were the most significant variable predicting an adaptive pattern of strategy use and learning attitudes. As expected, performance-approach goals were associated with an adaptive pattern of strategy use. Also as expected, extrinsic work goals did not predict significantly the use of learning and self-regulatory strategies. As for social enhancement goals, the current results showed clearly that these goals were maladaptive to learning. Overall, these results confirmed the relative importance of adaptive goals such as mastery-development goals and performance-

This study set out to examine the role of various goals and motivational beliefs on learning among distance learners. It adds to the distance learning literature by examining different goals that distance learners hold and the complex interaction relationship between

As expected, both efficacy and control beliefs were significant in predicting the use of learning and regulatory strategies, except for time management strategies. These results indicate that for this group of distance learners a strong sense of efficacy and control beliefs was associated with an engagement pattern that was characterised by adaptive strategy use. These two motivational beliefs also predicted distance learners' attitudes towards learning indicating that a stronger sense of efficacy and control beliefs was associated with learning enjoyment, interest and valuing of learning. To be motivated, distance learners need to feel

Another major aim of this study is to examine the moderation effects of efficacy and control beliefs on the impact of goals on strategy use and attitudes towards learning. This study found nonsignificant interaction effect between self-efficacy and various goals included in this investigation. In other words, a strong sense of efficacy beliefs would not enhance the positive effects of adaptive goals such as mastery and performance-approach goals. Neither would a strong sense of efficacy beliefs dampen the negative effects of less adaptive goals

It should be pointed out that contrary to Dweck's experimental study (Dweck, 1986; Elliot & Dweck, 1988), the current study examined distance learners' perceptions of learning at a rather general level. Dweck's study found significant interaction between self-efficacy and performance-approach goals on learning by requiring students in an experimental setting to complete clearly-defined academic tasks. Subsequent studies (e.g. Harackiewicz *et al.*, 2000; Kaplan & Midgley, 1997; Miller et al., 1993) using correlational design examining the moderation hypotheses at a rather general level did not found significant interaction between self-efficacy and performance goals on learning and achievement. The current

While the level of task specificity may be taken as an explanation for contradictory results between Dweck's study and the current study, a more critical factor may have been the

social goals did not show to have positive effects on learning attitudes.

approach goals on learning and attitudes among distance learners.

efficacious and certain about their learning progress.

such as extrinsic work goals and social enhancement goals.

nonsignificant results aligned with these correlational studies.

**5. Discussion**

motivational beliefs and goals.

salience of performance emphasis. Elliot and Dweck (1988) have created a high level of performance salience focusing participants to their perceived abilities to complete a specific experimental task. In this type of highly controlled setting, one's performance concerns and the demand of performance demonstration are heightened. Students' perceived level of efficacy will therefore be a crucial factor in moderating their performance goals on learning and achievement. Similarly, Church, Elliot & Gable (2001) and Braten and colleagues (2004) based on empirical evidence argued that the mediation role of self-efficacy with goals on learning will be more salient in a highly competitive and evaluative learning environment that focuses on relative performance. The participants and their learning context in the current study were radically different from these previous studies that used on-campus undergraduate students learning in a competitive environment or completing experimental tasks according to a set explicit performance criteria. Rather, the distance learning environment in this study promoted a mastery-focused orientation. Distance learners in this study engaged in their courses through distance learning mode in which face-to-face contact with other learners were limited to optional fortnightly tutorials. Distance learners are expected to complete the assigned readings and learning materials on their own, taking into consideration a suggested time-schedule for monitoring the progress themselves. In other words, the chance for distance learners to compete with each other was limited. Therefore, it can be argued that a lack of emphasis on relative performance might have led to the nonsignificant mediation effect of efficacy beliefs with goals, especially performanceapproach goals, on strategy use and learning attitudes.

Not only did the distance learning environment promoted mastery, the current sample of distance learners also held strong mastery goals ( *X* = 3.82). A mastery-focused learning system coupled with mastery-focused personal motivation would probably allow learners in this study to engage in adaptive patterns of strategy use and attitudinal development regardless of their level of efficacy beliefs (Dweck, 1986; Kaplan & Midgley, 1997). Kaplan and Midgley (1997, p.431) when concluding their study examining the moderation hypotheses stated that "*in an environment in which learning goals were emphasised more, level of perceived competence might become less influential for students with a predominant learning goal orientation*". In other words, mastery goals and mastery learning environment have predisposed distance learners to an adaptive pattern of strategy use and learning attitudes regardless their level of efficacy beliefs.

In contrast, the present study found several counts of significant interaction results between control beliefs, extrinsic work goals, and mastery goals. These significant results, though limited, confirmed the hypotheses set for the interaction between control beliefs and goals. These significant mediation effects indicate clearly that control beliefs will enhance the positive effects of adaptive goals such as mastery-development goals and dampen the negative effect of less adaptive goals, in this case, extrinsic work goals. These results suggest that control beliefs are critically important for distance learners. Within a distance learning system, distance learners are free to determine as to when they learn, how they learn and for what reasons they learn. A strong sense of control for distance learners means that they will be able to regulate their learning pace, determine the appropriate use of learning and regulatory strategies and adopt different goals for learning. This study showed that a strong sense of control beliefs enabled learners to use fewer surface strategies and more deep strategies even when holding extrinsic work goals that draw them away from engaging in the learning process. As for the case of mastery-development goals, a strong sense of control

The Role of Self-Efficacy, Control Beliefs and

Achievement Goals on Learning Among Distance Learners 247

certain extent, would be moderated by control beliefs. However these goals were not utterly detrimental to learning. Distance learners' career concerns were associated with a positive attitude towards learning. In other words, while a concern for career advancement will not associate learners with effective use of strategies, such a concern helps learners develop a positive attitude towards learning. Finally, social enhancement goals were found to have negative impact on learning as these goals were associated with surface strategies and the decrease in effort management. These associations seem to go against the Chinese collectivistic culture that places motivational significance on meeting social norms and fulfilling parental expectation on academic achievement. While it is not clear whether these social concerns would have positive impact on learners' achievement in this study, the present results indicate that a focus on enhancing social status and meeting social

expectation will draw learners away from meaningful engagement and persistence.

those focusing on various career considerations (Ng, 2008).

among distance learners.

**6. Conclusion**

Barron and Harackiewicz (2001) in a seminal paper argued that different achievement goals will have independent effects on learning and achievement, which can be assessed using the main effects of predictors in regression analyses. The current results clearly indicate that different goals had significant independent effects on learning and attitudes. Overall, the current findings suggest that goals that focus distance learners on the learning process are associated with adaptive use of strategies. In contrast, goals that draw distance learners away from the learning process and focus on the learning product do not predict the use of learning and regulatory strategies. Pintrich (2000) when discussing the nature of multiple goals argued that different goals will lead to different learning pathways and hence resulting in different learning experiences. The current results clearly support this notion of "multiple-goals, multiple-pathways". Distance learners focusing on mastery-development goals will certainly engage in and experience distance learning differently compared to

Barron and Harackiewicz (2001) also proposed that different goals may interact with each other. An additional regression analysis was conducted examining the interaction effects between different goals in this study. The result was nonsignificant, indicating that these goals did not interact with each other. In other words, a high level of performanceapproach goals would not enhance the positive effects of mastery-development goals. However, Ng (2008) using a clustering procedure found that distance learners are capable of endorsing a multitude of goals simultaneously. Future studies should continue to explore the potential interaction effects between goals on learning and engagement

This paper looked into the significant role of motivation in distance learning using an achievement goal framework. The results showed that conceptualising distance learners' motivation as motivational beliefs and goals adds to our understanding of distance learners' motivational and learning characteristics. Of course, the correlation nature of the current study does not allow for the analysis of causal ordering between beliefs, goals, strategies and attitudes. The culturally-specific and female-dominated sample prevents generalisation of the current findings. Further research should examine the relationship of these important variables using longitudinal design and include distance learners from other cultural contexts. The lack of achievement data was another major limitation in the current study.

beliefs simply enhanced the positive effects of these adaptive goals and led the learners to use fewer surface strategies.

Previous studies have examined the moderation effects using middle school students and young adolescents (e.g. Harackiewicz *et al.*, 2000; Kaplan & Midgley, 1997; Miller, Behrens, Greene & Newman, 1993). The current study provided empirical evidence on the moderation hypotheses among mature distance learners, which certainly added to our limited knowledge on the role of mediators on the effect of goals on learning. The present results focused us on the critical role of different moderators in distance learning. While the current study failed to find significant interaction between efficacy beliefs and goals on learning, the discussion above has highlighted the mastery-oriented nature of distance learning as an important factor that needs to be considered in examining the interaction effects. Certainly more research is needed to tease out the moderation effects of efficacy beliefs with goals among distance learners. In particular, our knowledge on extrinsic work goals and social enhancement goals are still rather limited. More research is required to understand how these goals operate in the learning process among distance learners.

The significant moderation effects of control beliefs reminded us the importance of considering additional mediators salient in a specific learning context. An experimental study conducted by Darnon and colleagues (2007) provided relevant empirical evidence showing that different forms of uncertainty created through negative feedback on performance or conflicting evaluation moderated the effects of performance-approach goals on learning. Within a distance learning system, tutor's disconfirming feedback on assessment may pose a significant threat to control beliefs (cf. Ng, 2009) and heighten a certain level of uncertainty. In addition, future research should extend the current investigation to cover different type of dependent measures and examine the moderation hypotheses within different level of task specificity. Two important dependent measures are distance learners' persistence and achievement levels. Will strong efficacy or control beliefs act as a buffer against the negative impact of less adaptive goals such as extrinsic work goals on persistence and achievement? In addition, the current study focused on learners' motivation and learning at the course level without referring to specific academic tasks. Future studies should consider the moderation hypotheses using more specific academic tasks such as sitting for an end-of-course examination that highlights performance demonstration and the critical importance of efficacy and control beliefs.

With regard to the predictive importance of goals, the regression models consistent with past achievement goal research showing that mastery-development goals, and to a lesser extent performance-approach goals, were associated with adaptive patterns of strategy use and learning attitudes. These results were achieved after controlling for the effects of both efficacy and control beliefs. The significance of mastery-development goals was demonstrated in the associated engaged pattern characterising by adaptive use of strategies. The use of regulatory strategies, and more specifically, effort management strategies indicate that distance learners focusing on mastery and development goals will be more likely to persist and continue investing effort on learning despite obstacles and problems. Performance-approach goals in this study were also positively related to these strategies. In addition these goals focused learners on time management and organisation. Extrinsic work goals did not predict any of the strategies. The interaction results indicated that extrinsic work goals were probably associated with a maladaptive pattern of strategy use, which to a

beliefs simply enhanced the positive effects of these adaptive goals and led the learners to

Previous studies have examined the moderation effects using middle school students and young adolescents (e.g. Harackiewicz *et al.*, 2000; Kaplan & Midgley, 1997; Miller, Behrens, Greene & Newman, 1993). The current study provided empirical evidence on the moderation hypotheses among mature distance learners, which certainly added to our limited knowledge on the role of mediators on the effect of goals on learning. The present results focused us on the critical role of different moderators in distance learning. While the current study failed to find significant interaction between efficacy beliefs and goals on learning, the discussion above has highlighted the mastery-oriented nature of distance learning as an important factor that needs to be considered in examining the interaction effects. Certainly more research is needed to tease out the moderation effects of efficacy beliefs with goals among distance learners. In particular, our knowledge on extrinsic work goals and social enhancement goals are still rather limited. More research is required to

understand how these goals operate in the learning process among distance learners.

demonstration and the critical importance of efficacy and control beliefs.

The significant moderation effects of control beliefs reminded us the importance of considering additional mediators salient in a specific learning context. An experimental study conducted by Darnon and colleagues (2007) provided relevant empirical evidence showing that different forms of uncertainty created through negative feedback on performance or conflicting evaluation moderated the effects of performance-approach goals on learning. Within a distance learning system, tutor's disconfirming feedback on assessment may pose a significant threat to control beliefs (cf. Ng, 2009) and heighten a certain level of uncertainty. In addition, future research should extend the current investigation to cover different type of dependent measures and examine the moderation hypotheses within different level of task specificity. Two important dependent measures are distance learners' persistence and achievement levels. Will strong efficacy or control beliefs act as a buffer against the negative impact of less adaptive goals such as extrinsic work goals on persistence and achievement? In addition, the current study focused on learners' motivation and learning at the course level without referring to specific academic tasks. Future studies should consider the moderation hypotheses using more specific academic tasks such as sitting for an end-of-course examination that highlights performance

With regard to the predictive importance of goals, the regression models consistent with past achievement goal research showing that mastery-development goals, and to a lesser extent performance-approach goals, were associated with adaptive patterns of strategy use and learning attitudes. These results were achieved after controlling for the effects of both efficacy and control beliefs. The significance of mastery-development goals was demonstrated in the associated engaged pattern characterising by adaptive use of strategies. The use of regulatory strategies, and more specifically, effort management strategies indicate that distance learners focusing on mastery and development goals will be more likely to persist and continue investing effort on learning despite obstacles and problems. Performance-approach goals in this study were also positively related to these strategies. In addition these goals focused learners on time management and organisation. Extrinsic work goals did not predict any of the strategies. The interaction results indicated that extrinsic work goals were probably associated with a maladaptive pattern of strategy use, which to a

use fewer surface strategies.

certain extent, would be moderated by control beliefs. However these goals were not utterly detrimental to learning. Distance learners' career concerns were associated with a positive attitude towards learning. In other words, while a concern for career advancement will not associate learners with effective use of strategies, such a concern helps learners develop a positive attitude towards learning. Finally, social enhancement goals were found to have negative impact on learning as these goals were associated with surface strategies and the decrease in effort management. These associations seem to go against the Chinese collectivistic culture that places motivational significance on meeting social norms and fulfilling parental expectation on academic achievement. While it is not clear whether these social concerns would have positive impact on learners' achievement in this study, the present results indicate that a focus on enhancing social status and meeting social expectation will draw learners away from meaningful engagement and persistence.

Barron and Harackiewicz (2001) in a seminal paper argued that different achievement goals will have independent effects on learning and achievement, which can be assessed using the main effects of predictors in regression analyses. The current results clearly indicate that different goals had significant independent effects on learning and attitudes. Overall, the current findings suggest that goals that focus distance learners on the learning process are associated with adaptive use of strategies. In contrast, goals that draw distance learners away from the learning process and focus on the learning product do not predict the use of learning and regulatory strategies. Pintrich (2000) when discussing the nature of multiple goals argued that different goals will lead to different learning pathways and hence resulting in different learning experiences. The current results clearly support this notion of "multiple-goals, multiple-pathways". Distance learners focusing on mastery-development goals will certainly engage in and experience distance learning differently compared to those focusing on various career considerations (Ng, 2008).

Barron and Harackiewicz (2001) also proposed that different goals may interact with each other. An additional regression analysis was conducted examining the interaction effects between different goals in this study. The result was nonsignificant, indicating that these goals did not interact with each other. In other words, a high level of performanceapproach goals would not enhance the positive effects of mastery-development goals. However, Ng (2008) using a clustering procedure found that distance learners are capable of endorsing a multitude of goals simultaneously. Future studies should continue to explore the potential interaction effects between goals on learning and engagement among distance learners.


**2. The over-simplification of classroom teaching and learning**

There is widespread support in the literature for the view that primary classroom learning and teaching are not straightforward processes. Davis and Sumara [1] note that most teachers will attest to the unpredictability of learners' responses to teaching. Eisner [2] described teaching as "an inordinately complex affair" and others, [3–5] have framed teachers' roles in terms of managing uncertainty and problematizing unpredictability. Shulman [6] is unequivocal in describing teaching as "perhaps the most complex, most challenging, and most demanding, subtle, nuanced and frightening activity that our species ever invented". Descriptions of the facilitation and elicitation of learning by teachers themselves [7, 8] also acknowledge the unpredictable, dynamic and often messy, uneven nature of learning, and Alexander et al. sum up the argument with the assertion that "one cannot begin to understand the true nature of human learning without embracing its interactional complexity [9]." The case for learning and teaching being far from straightforward is also captured succinctly by Schon, who described teachers' work as operating in the "swampy lowlands" of everyday life. For him, "the problems of real-world practice do not present themselves to practitioners as well-formed structures. Indeed, they tend not to present themselves as problems at all but as messy indeterminate situations [10]." Considering this, it seems reasonable to suggest that what pupils learn in the context of school classrooms does not flow mechanistically from teachers' input and is not entirely within the conscious control of either teachers or pupils. This is not to argue that learning is not a function of teaching, but that pupil learning must be driven by more than merely the influence of teachers and teaching. It seems reasonable to hypothesize that teaching input is filtered through a range of networked factors resulting in learning occurring sometimes because of, sometimes despite, and sometimes irrespective of what teachers do.

Notwithstanding these depictions of the complex nature of learning and teaching, including from teachers themselves, a popular portrayal in policy and public discourse runs counter to this, presenting teaching and learning as simple, linear, causal, and mechanistic activities [11] reflecting a technical rationalist view of the profession [12, 13]. In this conception, teachers simply apply instrumental "teaching" solutions to address well-formed "learning" problems. Discourse and national agendas concerning teaching, learning, pupil progress, curriculum, standards, and teacher professional development are typically driven by this input-output conception. In the dominant policy discourse, "outstanding" teaching is often narrowly defined as the meticulous planning of lessons to meet specific, predetermined objectives [14] and despite years of reform in the UK and comparable education systems, the language of policy [15] still partially depicts a transmission and absorption notion of teaching and learning. This leads to the popular notion that if teaching is "outstanding" learning will (or should) be too. Tessellating policies of national testing, league tables, school inspection, and teacher competency descriptors firmly position teachers as the lynchpins [16] of pupil progress with the consequence that they are routinely held accountable for a phenomenon (learning) which appears to be only partially within their control.

This technical rationalist positioning of teaching alludes to an absorption and output conception of learning and learners. However, the pupil end of any learning and teaching relationship is no less complex than the teaching end. In any given school classroom "the persistence of inequities in student achievement [17]" speaks to a range of factors influencing learning. These include inherited and environmental

#### *Classroom as Complex Adaptive System and the Emergence of Learning DOI: http://dx.doi.org/10.5772/intechopen.101699*

predispositions such as cognitive ability, personality, confidence, task commitment, and risk-taking tendencies along with influences from home and school ecosystems. More ephemeral factors influencing volition such as social dynamics, nutrition, mood, and even weather may also play a part. Research from several paradigms offers insights into learner factors and their effects on learning outcomes. Bronfenbrenner's [18] Ecological systems theory, for example, presents a framework for describing community and environmental influences on individuals. According to the model, interaction with other individuals and institutions across five levels of systems (from micro to chrono) shapes the individual's growth, learning, and development. These include family, peer group, school, media, and health care policies for example. Research into the emergence of gifts and talents in school-age learners [19, 20] has highlighted common elements, which typically correlate with high performance, including general cognitive ability, environment, personality, self-confidence, and chance. Numerous studies from the field of psychology [21–24] illustrate how personality influences readiness to learn and learning outcomes. Common findings suggest that the long-established "Big Five" personality traits (openness, conscientiousness, extraversion, agreeableness, and neuroticism) interact in statistically significant ways with learning. Studies of class emotion and mood [25–27] reveal how pupil interactions, on/off-task behaviors, and learning can be influenced by interpersonal features including regulation, negotiation, and resistance. Despite wide-ranging acknowledgment across multiple disciplines that learning is influenced by a complex array of converging and mutually interacting factors (those mentioned above and many not mentioned e.g. working memory, self-efficacy, parents' education, personal health, and cultural expectations to name a few), its depiction remains largely characterized in public discourse by the receiving and remembering of information and the mastery of a set of skills. This is evidenced most clearly in the way that policy developments in the United Kingdom, and comparable education systems, over the last 20 years have striven to routinize teaching.

It is against this landscape, in which teaching and learning are characterized quite differently in policy and media to how teachers and pupils experience them, and how research frames them, that this discussion chapter sits. I have previously suggested [28] there is a need for more accurate depictions of teachers' work and its relationship to pupils' learning. As Hardman [29] points out, there has been a failure of simple causal explanations to adequately account for the complexities of school and classroom learning, because, according to Davis & Sumara learning tends to "defy simplistic analyses and cause-effect explanations [1]". Complex adaptive systems (CAS) theories may offer valuable insights about ways that learning emerges in classrooms and my hope is that advancing this discussion will contribute to unpicking current over-simplified thinking about teaching and learning and offer new perspectives on pedagogy.