The realisation of integrated, optimised, multi-nodal networks featuring urban rail within densely populated, commercialised, and industrialised areas, and regional rail within larger agglomerations and corridors, is already visible in several countries. Consider the example of Verkehrsverbund Rhein-Ruhr in Germany, serving a conurbation of eight million inhabitants in a 7,300 km2 area. Formerly isolated light rail networks of individual local authorities have been connected to one another, going so far as to regauge some from metre gauge to standard gauge to promote networkability. This has in turn made acquisition of new 160 km/h regional trains sensible, to reduce journey times over longer distances. At national and international scale, long distance, high-speed passenger and heavy freight trains are common in Europe and North America, respectively. Dedicated freight and passenger corridors, where traffic volume is sufficient to separate freight and passenger trains, have been realised in China and India. China's One Belt, One Road framework takes rail-based corridor development to an unprece‐ dented level. So the building blocks of the Railway Enlightenment or New Normal exist: What should now follow is expansion of application to countries that are earnest about road-to-rail shift to reduce congestion and the dependence on fossil fuels and the accompanying pollutant emissions.

## **4.4. Further research**

England. Railways have settled into a new dispensation. From a position of strength, their abilities stretch from heavy-duty urban transport across continents to distant strategic

The 14th–17th century Renaissance, or rebirth, provides a fitting archetype of the 1964–1989 quarter-century railway renaissance. Looking back to 1989, it is evident that the following quarter century to 2014 has been a period of settling railways into their four inherently competitive sub-modes high-speed intercity, heavy haul, heavy intermodal, and contempo‐ rary urban rail. The Enlightenment, the period that followed the Renaissance and exploited

The know-how of the railway renaissance has been captured in a myriad of publications and widely dispersed railway industry institutional knowledge. Now freely available, it is being applied to position railways for an energy-scarce future. Global consensus seems to be building around the year 2050 as the date by which the shift from a fossil-fuelled economy to a renewables-powered economy should be largely in place. The timing recognises the need to wear out the present fleet of transport vehicles—planes, ships, and trains—and to adapt, construct, or renew infrastructure and rolling stock. The exact year is not important. What is important is the form that transport systems will take at that time. The European Commission's [10] Transport White Paper, for example, provides valuable

**•** An efficient core network for multimodal intercity travel and transport with greater use of buses and coaches, rail and air transport for passengers and, for freight, multimodal

**•** New transport patterns emerging, according to which larger volumes of freight and greater numbers of travellers are carried jointly to their destination by the most efficient (combina‐

**•** A global level playing field for long-distance travel and intercontinental freight in which

**•** Clean urban transport and commuting where a higher share of travel is by collective transport, and the interface between long distance and last-mile freight transport is organ‐

The realisation of integrated, optimised, multi-nodal networks featuring urban rail within densely populated, commercialised, and industrialised areas, and regional rail within larger agglomerations and corridors, is already visible in several countries. Consider the example of Verkehrsverbund Rhein-Ruhr in Germany, serving a conurbation of eight million inhabitants in a 7,300 km2 area. Formerly isolated light rail networks of individual local authorities have been connected to one another, going so far as to regauge some from metre gauge to standard gauge to promote networkability. This has in turn made acquisition of new 160 km/h regional trains sensible, to reduce journey times over longer distances. At national and international scale, long distance, high-speed passenger and heavy freight trains are common in Europe and North America, respectively. Dedicated freight and passenger corridors, where traffic volume

the intellectual freedom that it had let loose, has inspired this section.

26 Railway Research - Selected Topics on Development, Safety and Technology

solutions relying on waterborne and rail modes for long-hauls.

(high speed) rail should absorb much medium distance traffic.

perspectives on that time by envisioning:

tion of) modes.

ised more efficiently.

horizons.

Despite the difficulty of obtaining requisite data, the authors, nevertheless, continue seeking substitute sources that could support multivariate statistical analysis of post renaissance railway positioning. Substantial movement within the clusters is evident, so it is expected that changes in the composition of the variables used, and of course a different time period, could possibly identify new, more refined factors and clusters. Having established the benefits of achieving renaissance, it would now be useful to understand how to further optimise the role of railways in societies that are progressing well in their transition to an energy-scarce future.

## **5. Conclusions**

For convenience, the research question repeated from section 2.1 is: *Does a new post-renaissance normal now prevail, and as a corollary, does there seem to be any viable alternative*?

Rail is to be observed doing the transport heavy lifting in countries that have embraced railway renaissance, as it should do by virtue of strong positioning in its inherently competitive market spaces and its comparative energy efficiency. Transcontinental and intercontinental hauls at higher speed and lower energy consumption than waterborne transport are emerging. Regional rail is expanding around the world, frequently with double-deck trains. High speed finds favour as an effective economic and social development intervention. Urban rail is standardising on standard gauge, even in non-standard gauge countries such as Brazil and India, to economically deliver high capacity in burgeoning cities. Urban guided transit scales capacity offerings from rubber-tyred light solutions for small cities to heavyweight metro solutions for densely populated cities. No solution other than heavy haul comes to mind to move large tonnages of bulk minerals over long distances. Double-stack container trains are able to wrest high-value, low-density goods from road hauliers. Overall, railway renaissance has aligned customers and suppliers around what works and what does not work in a global market, enabling suppliers to produce widely acceptable products and solutions at competitive prices.

It is now evident that the answer to the research question is that countries that have embraced railway renaissance have been able to reposition their railways to play a substantial role in their economies and societies. In the sense that the Enlightenment exploited the intellectual freedom that it had let loose, yes, that is happening today in the post railway renaissance period. In present colloquial language, the Enlightenment would more likely be called the New Normal. And that is just what has happened over the five decades since 1964. It is once again business as usual, the material difference being that the rail mode is a now a formidable contender in the High Speed, Heavy Haul, Heavy Intermodal and Urban Rail market spaces. The authors therefore argue that the hypothesis has not been disproved.

Regarding the corollary to the research question, there is no evidence of railways emerging from the Fortuitous an Insecure clusters other than by following one or more of the renaissance growth curves through appropriate investment.

## **Author details**

Dave van der Meulen\* and Fienie Möller

\*Address all correspondence to<EMAIL_ADDRESS>
Railway Corporate Strategy CC, South Africa

## **References**

nal of Rail and Rapid Transit, 228(6), a Special Issue from the 2013 International Heavy Haul Association Conference. 2014.

[8] Garcia A. High speed, energy consumption and emissions. Paris: International Union of Railways. 2010.

business as usual, the material difference being that the rail mode is a now a formidable contender in the High Speed, Heavy Haul, Heavy Intermodal and Urban Rail market spaces.

Regarding the corollary to the research question, there is no evidence of railways emerging from the Fortuitous an Insecure clusters other than by following one or more of the renaissance

[1] Van der Meulen RD, Möller LC. Railway globalization: Leveraging insight from de‐ veloped- into developing regions. Proceedings of the 7th World Congress on Railway

[2] Van der Meulen RD, Möller LC. Ultimate interoperability: Line-haul railways as global corporate citizens. Proceedings of the 8th World Congress on Railway Re‐

[3] Van der Meulen RD, Möller LC. Competitiveness and sustainability of railways. In X. Perpiñà (Ed.): Infrastructure design, signalling and security in railway. Rijeka, Cro‐

[4] Van der Meulen RD, Möller LC. European- and global urban guided transit: Greenand socio-economic fit. Proceedings of the Transport Research Arena, Procedia – So‐ cial and Behavioral Sciences 48. Athens, Greece: Programme Committee of the

[5] Van der Meulen RD, Möller LC. Towards sustainable heavy haul traction energy: A review. Proceedings of the 10th International Heavy Haul Conference. New Delhi,

[6] Van der Meulen RD. From global research to national rail policy. Proceedings of the Transport Research Arena, Outreach Marketplace Poster #28020. Paris, France: Pro‐

[7] Van der Meulen RD, Möller LC. Sustainable heavy haul traction energy: A review of systemic issues. Proceedings of the Institution of Mechanical Engineers, Part F: Jour‐

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growth curves through appropriate investment.

28 Railway Research - Selected Topics on Development, Safety and Technology

**Author details**

**References**

Dave van der Meulen\*

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[26] Des Longchamps H. World Bank involvement in the railways sector in Sub-Saharan Africa. In 3rd Annual African Rail Summit, 18-19 November 2014. Johannesburg,

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[29] Kurosaki F, Okuda K. On-rail competition in Korea: A comparison with railways in Japan and Europe. Proceedings of the Eastern Asia Society for Transport Studies, 9. 2013. http://easts.info/on-line/proceedings/vol9/PDF/P89.pdf (accessed 3 May 2014).

[30] European Rail Research Advisory Council. Freight Roadmap 2012. http:// demo.oxalis.be/errac/errac\_website/wp-content/uploads/2013/06/errac\_freight\_road‐

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[34] Lötschberg Base Tunnel. http://en.wikipedia.org/wiki/L%C3%B6tschberg\_Base\_Tun‐

[35] Bullock R. Results of Railway Privatization in Africa. http://siteresources.world‐ bank.org/EXTRAILWAYS/Resources/515244-1268663980770/privatization\_africa.pdf

[36] Van der Meulen RD. Global railway renaissance and its influences on logistics and mobility in Africa. Africa Forum 2015. Arusha, Tanzania: Chartered Institute of Lo‐

[37] Wang L. China's High-speed rail diplomacy. http://www.worldcrunch.com/rss/ world-affairs/china-039-s-high-speed-rail-diplomacy/railways-high-speed-markettechnology-li-keqiang/c1s16312/#.VOHHSmccR-Y (accessed 16 February 2015).

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**8. Immunosuppression**

• Cross-clamping of the supraceliac aorta is performed simultaneously with or immediately

• In situ cooling of abdominal organs, and exsanguination before removing the organs to the

• University of Wisconsin (UW) Universal Organ Preservation solution for both in situ flush-

• Retrieve iliac, brachiocephalic and/or carotid arteries and veins as potential vascular grafts.

Intestinal grafts require minimal back table; however, this depends on the retrieval technique and the extension of the allograft. Most commonly, back table involves to identify and tie the lymphatics. If the pancreas is retrieved along with a small bowel only graft, then it has to be

• Implantation begins commonly with adhesiolysis, as adhesions are usually abundant sec-

• Depending on the type of transplant, the aorta and IVC or SMA and SMV/PV are dissected

• Venous anastomosis to the graft is frequently performed to the recipient cava. However,

• After reperfusion of the graft and careful hemostasis, the proximal and distal ends of the intestinal graft are anastomosed to the proximal and distal ends of the native digestive

• Most centers bring out a temporary stoma by utilizing various techniques (e.g. Bishop Koop), for post-operative endoscopic surveillance. This stoma is usually reversed in

• Closure of the abdominal wall can be very challenging and should not be attempted under tension; if this is the case, keeping the abdominal wall open and planning for a sequential closing is preferable. Some centers are routinely performing abdominal wall transplanta-

Patients are monitored in intensive therapy unit (ITU) post-operatively. It is common practice to administer broad-spectrum intravenous antibiotics and antifungals for 5–7 days post transplantation. Blood tests are sent daily and as well as arterial/venous blood gases to check

• The proximal and distal ends of the native digestive track are identified and dissected.

• Arterial anastomosis is performed to the abdominal aorta via arterial jump graft.

when possible, venous anastomosis to the portal system is preferred.

track. In some cases, the distal end is brought out as a permanent stoma.

tion from the same deceased donor in order to achieve closure.

following venting of the IVC or atrium and cold perfusion is commenced.

back table for preparation.

**6.2. Back-table preparation of organs**

removed/sacrificed on the back table.

ondary to previous surgeries.

and mobilized for the vascular anastomosis.

**6.3. Recipient operation**

6–12 months.

**7. Postoperative considerations**

bleeding and homeostasis.

ing and cold storage is most frequently used.

298 Organ Donation and Transplantation - Current Status and Future Challenges

The intestine is the largest lymphoid organ in the body and hence, appropriate immunosuppression has been a real challenge. The lack of effective immunosuppressive agents hampered the first attempts of ITx in the 1960s. Over the years, advances in immunosuppression have transformed ITx with the intent of attenuating the intestinal allograft immunity and shifting it to a tolerogenic status [32].

Induction strategies to minimize rejection by reducing the recipient's T-cell load were implemented, initially with cyclophosphamide induction therapy, which was later replaced by daclizumab, an interleukin-2 receptor antagonist (IL2RA) [33]. Basiliximab, another IL2RA, in addition to tacrolimus and prednisone immunosuppression has also been utilized and shown to decrease the incidence of acute rejection [34, 35].

Alemtuzumab induction is becoming increasingly popular and Lauro et al. [36] reported significantly less early rejection episodes, with no sepsis implications. The use of Basiliximab monthly as part of maintenance immunosuppression has been associated with a decrease in acute rejection in liver-excluding transplants [37].

Immunosuppression regimen varies, with several protocols having been reported: Tacrolimus and steroids (35.8%) followed by tacrolimus, mycophenolate and steroids (18.7%), tacrolimus and mycophenolate without steroids in 15.4% of cases and tacrolimus alone in 13.8% of cases [38].

Most centers will endeavor to perform early, frequent endoscopies because of high prevalence of acute rejection in the early post transplantation phase [21] and then continue with

Intestinal Transplantation

301

http://dx.doi.org/10.5772/intechopen.74825

The Oxford group, who is utilizing vascularized composite allograft (VCA) transplants from the same donor [45], is now mostly relying on the VCA as a surrogate marker for rejection and is not strictly adherent to early, intensive intestinal biopsy protocols [46]. They have reported that the VCA can provide lead time (about 7 days) before the onset of bowel dysfunction and

Rejection episodes are usually treated with pulses of methylprednisolone, and in resistant

A recent case series reported good outcomes in ITx of positive cross-match patients with only one patient developing acute rejection, with the use of intravenous immunoglobulin (IVIG) and rituximab [47]. Bortezomib, a proteasome inhibitor, has also been recently shown to be effective against donor-specific antibody (DSA)-related rejection [48]. Recently, Eculizumab, a monoclonal antibody that inhibits complement factor 5a, was shown to be effective in maintaining a 2-year rejection-free period in a highly sensitized patient [49]; however, high costs

As mentioned before, biomarkers such as citrulline have gained interest in recent years. The prospective cohort study by Hibi et al. [50] reported excellent negative predictive value (range 93–99%) for citrulline levels as exclusionary marker for all types of acute rejection (cut-off point, 20 mmol/l) and moderate or severe acute cellular rejection (cut-off point, 10 mmol/l). Another study by the Bologna group [51] showed that citrulline sensitivity and specificity in detecting acute rejection, when adjusted for chronic renal failure, almost doubled the sensitivity of citrulline as a non-invasive marker of acute rejection in ITx. In general, citrulline can act as an exclusionary tool, as high levels are unlikely to be found in intestinal allograft rejection. Recently, a large series evaluating video capsule endoscopy has shown to be potentially useful in the diagnostic management of patients with ITx [52]. Other non-invasive predictors of rejection include a recent retrospective study that revealed low insulin-like growth factor-1 and high calprotectin levels during malnutrition states post-ITx, and these findings should prompt the clinicians to investigate for acute rejection or infection [53]. Circulating apoptotic T cells following Caspase 3 activation may be a non-invasive marker for patients who are less

Chronic rejection is diagnosed histologically with the identification of an obliterative arteriopathy in medium-sized vessels in the serosal layer with diffuse concentric intimal thickening [55]. This necessitates full-thickness biopsy and makes diagnosis challenging. Chronic rejection is clinically associated with diarrhea, ulceration, focal loss of mucosal folds, mural thickening and pruning of mesenteric artery arcades [55]. Surgery in such a hostile environment may lead to unwanted enterotomies and fistulae. Re-transplantation should therefore

likely to have rejection episodes than those who have lower levels [54].

regular endoscopies as part of their surveillance protocol [44].

this could be proven as a unique prognostic tool [45].

cases, thymoglobulin [32] or alemtuzumab [46].

make this approach prohibitive for general use.

**9.2. Chronic rejection**

Target trough levels of tacrolimus vary between centers. Pittsburgh has reported target levels between 10 and 15 ng/ml in the first 3 months and thereafter 5–10 ng/ml [39]. Tacrolimus with low-dose steroids remains the most effective and durable long-term combination therapy [21] and is the most common maintenance immunosuppressive regimen [7].

Sirolimus, a rapamycin inhibitor, has been shown as a useful adjunct to tacrolimus in the presence of nephrotoxicity or rejection [40]. However, it carries the disadvantage of severe debilitating oromucosal ulceration. Azathioprine and mycophenolate mofetil have also been used as adjunctive immunosuppressive therapies [33]. Mycophenolate mofetil, however, causes symptomatic diarrhea (increased stoma output) and microscopically evident apoptosis in approximately 40% of solid organ transplant recipients, which could regrettably be mistaken for rejection [41].


2.2.4. Novel therapeutic strategies against MDSCs

Although various agents such as chemotherapeutic drugs, vitamin derivative, and mAb are widely applied against MDSCs, only few of them are known to exhibit their inhibitory effect directly on MDSCs. Moreover, restricted use of these drugs on certain conditions or cancer patients often opens the possibility to apply emerging agents that bear direct inhibitory effect on MDSCs.

Earlier investigations reveal CuNG as a strong immunomodulator that successfully induce the conversion of immunosuppressive TAMs toward proimmunogenic type, enhancing Th1 response sufficient to resolve drug-resistant cancer in animal model. Interestingly, the effect of CuNG is not exclusive to TAMs; rather it also targets MDSCs which happens to be another strong negative regulator of tumor immunity [55]. Treatment with CuNG in drug-resistant mice severely abrogates the accumulation of MDSCs in spleen as well as tumor site. Further study discloses that this reduction was associated with enhanced Th1 responses (evident from increase in IFN-γ production) and diminished Th2 responses (decrease in IL-4 production) as well as decline in Foxp3+ Treg numbers [55]. Detailed investigation reveals the underlying mechanism that is associated with Fas-FasL interaction between MDSCs and CD4+ T cells. Treatment of CuNG increases the FasR expression in MDSCs and FasL in CD4+ T cells. Although FasL expression is enhanced, FasR expression in CD4+ T cells was not observed following the treatment that determines the specificity of CuNG [55].

Another study that deals with metal chelate of Schiff's base is associated with the immunomodulation of tumor microenvironment. Metal chelate of manganese (MnNG) can have a number of valence states and possess the ability to modulate the immunosuppressive TME by reducing the number of tumor-associated MDSCs in drug-resistant tumor-bearing mice. A closer look at MnNG-mediated action reveals that it promotes MDSC differentiation into dendritic cells but not toward macrophages [56]. Additionally, it helps in the proliferation of T cells that ultimately skew Th2 immunity toward protective Th1 response. Thus MnNG may help in the differentiation of immunosuppressive MDSCs toward dendritic cells and with the upregulation of costimulatory molecules, namely CD80 and CD86, enabling them to act as better antigenpresenting cells (APCs) that ultimately generate protective antitumor immunity [56]. However, the plausible molecular mechanism underlying MnNG-mediated differentiation of MDSCs in TME is still elusive and warrants further investigations.

## 2.3. T-regulatory cells in cancer

led to MDSC-derived IL-1β secretion and angiogenesis [53]. Attempts have also been made to deplete Gr-1+ MDSCs by using anti-Gr-1 antibody that result in delayed tumor growth in mice [53]. However, this antibody also targets neutrophils, thus lacking the necessary specificity for

Fourthly, another attractive way to control MDSC function would be interrupting the underlying signaling pathways that are responsible for the production of suppressive factors by these cells. The molecules that can be effectively targeted for this purpose include cyclooxygenase 2, arginase 1, iNOS, and indoleamine 2,3-dioxygenase [52]. Since ARG1 and iNOS are the primary enzymes responsible for MDSC immunosuppression, these enzymes are the most likely targets for novel therapeutic interventions. Various different drugs including nitroaspirin, COX-2 inhibitors, and phosphodiesterase-5 (PDE5) inhibitors have been shown to profoundly inhibit both ARG1 and iNOS activity in MDSC. By removing MDSC suppressive mediators, these drugs exhibited a potent ability to restore antitumor immune responses and delayed tumor progression in several mouse models [52–54]. Interestingly, in addition to inhibiting MDSC function, COX2 inhibitors also blocked the systemic development of MDSC as well as CCL2-mediated accumulation of these cells in the tumor microenvironment in a

Although various agents such as chemotherapeutic drugs, vitamin derivative, and mAb are widely applied against MDSCs, only few of them are known to exhibit their inhibitory effect directly on MDSCs. Moreover, restricted use of these drugs on certain conditions or cancer patients often opens the possibility to apply emerging agents that bear direct inhibitory effect

Earlier investigations reveal CuNG as a strong immunomodulator that successfully induce the conversion of immunosuppressive TAMs toward proimmunogenic type, enhancing Th1 response sufficient to resolve drug-resistant cancer in animal model. Interestingly, the effect of CuNG is not exclusive to TAMs; rather it also targets MDSCs which happens to be another strong negative regulator of tumor immunity [55]. Treatment with CuNG in drug-resistant mice severely abrogates the accumulation of MDSCs in spleen as well as tumor site. Further study discloses that this reduction was associated with enhanced Th1 responses (evident from increase in IFN-γ production) and diminished Th2 responses (decrease in IL-4 production) as

mechanism that is associated with Fas-FasL interaction between MDSCs and CD4+

Treatment of CuNG increases the FasR expression in MDSCs and FasL in CD4+

following the treatment that determines the specificity of CuNG [55].

Although FasL expression is enhanced, FasR expression in CD4+ T cells was not observed

Another study that deals with metal chelate of Schiff's base is associated with the immunomodulation of tumor microenvironment. Metal chelate of manganese (MnNG) can have a number of valence states and possess the ability to modulate the immunosuppressive TME by reducing the number of tumor-associated MDSCs in drug-resistant tumor-bearing mice. A closer look at MnNG-mediated action reveals that it promotes MDSC differentiation into dendritic cells but not toward macrophages [56]. Additionally, it helps in the proliferation of T cells that ultimately

Treg numbers [55]. Detailed investigation reveals the underlying

T cells.

T cells.

clinical use.

on MDSCs.

mouse model of glioma [52, 54].

178 Anti-cancer Drugs - Nature, Synthesis and Cell

well as decline in Foxp3+

2.2.4. Novel therapeutic strategies against MDSCs

The tumor escape of immune surveillance is also achieved by the recruitment of immunosuppressive CD4+ CD25+ FoxP3+ Tregs into the tumor microenvironment. The phenotype and functional properties of Tregs are similar in mice and human. They include both natural Treg (nTreg) and locally induced Treg (iTreg) cells [57, 58]. nTreg cells are derived from thymus without specific antigenic stimulation and represents a small fraction (5–6%) of total CD4+ T cells. They are characterized by the expression of CD25, FoxP3, and GITR [3]. CD4+ or IL-10 expressing iTreg cells are also known to express high levels of Foxp3 and GITR [58]. Induced Tregs are emerged from naive T cells by specific modes of antigenic stimulation, especially in a particular cytokine milieu [57, 58].  A large number of CD4+ CD25+ FoxP3+ Treg cells are found in both the circulation and tumor site of different cancer-bearing patients and their number inversely proportional to the survival of the patients. They are either naturally occurring or locally induced Treg population [58]. Experimental observation reveals that chemokine CCL2 produced by tumor and its associated macrophages facilitate the recruitment of thymus-derived Treg cells in tumor microenvironment [3]. Defective myeloid DCs also induced IL-10+ regulatory T cells in vitro and in vivo in patients with cancer. Conversion of Treg from CD4+ CD25− T cells depends on local cytokine milieu. Immunosuppressive cytokines such as TGF-β and IL-10, which are present at high levels in the tumor microenvironment, might mediate induction and differentiation of iTreg cells [58]. Recent studies also highlighted the role of MDSCs to facilitate the de novo development of Tregs through TGF-β dependent and independent pathways [34].

#### 2.3.1. Tregs and immunosuppression

Tumor-associated Tregs suppress both the innate and the adaptive immune responses. The suppressive function of tumor-induced Tregs depends on the antigen-specific stimulation of these cells. Once activated, they efficiently suppress CD4+ and CD8+ T cells in an antigen nonspecific manner [58]. This suppressive potential of Tregs depends on the cell-cell contact and soluble mediators (e.g., TGF-β and IL-10) released by Tregs. Recent evidence suggests that Treg exerts immunosuppressive function by lowering the endogenous TAA-specific T-cell immunity, thereby contributing to tumor progression [3]. Furthermore, the progressive infiltration and induction of Treg tilt the balance between Treg cells and effector T cells toward immunosuppression. This is, in part, mediated by competition for IL-2 between Tregs and conventional T cells. IL-2 promotes the differentiation of T cells into effector T cells. Interestingly, it also promotes the differentiation of immature T cells into regulatory T cells. Therefore, competitive consumption of IL-2 is also indirectly related to suppressive mechanism for Tregs in established tumors [59]. Another effective way to suppress T-cell activation and promote tolerance is to reduce the availability of tryptophan, which is performed by the IDO+ APC in the tumor microenvironment. CTLA4+ Tregs help inducing the expression of IDO in APCs through CTLA4 to mediate their suppressive activity [60]. Furthermore, Tregs also downregulate the expression of CD80 and CD86 through CTLA-4 and induce immunosuppressive costimulatory molecule B7-H4 on DCs by IL-10 and thus hamper activation of other T cells by DCs [61]. Additionally, Tregmediated suppression of natural killer (NK)-cell function is associated with TGF-β in tumorbearing mice [62].


3.2.3. Discussion

When taken in conjunction with earlier findings [3,43,44], the distribution of function words we observed here supports the suggestion that they form a natural communicative distribution. This in turn suggests that, despite the fact that prepositions (in contrast to determiners and pronouns) distinguish between spatial and temporal relations, prepositions, determiners, and pronouns are part of the same functional subsystem and, at some level, serve the same communicative function.

By contrast, we find that the lexically more diverse categories fit power laws. As previously discussed, these distributions could be the product of aggregating over multiple communicative distributions serving distinct communicative functions. This suggestion is further supported by the observed distributions of verbs and nouns, which suggest that a smaller number of unique verb types appears across a larger number of distinct communicative contexts than is the case for nouns. This observation is supported by the fast growing head in the verb distributions, which appears to result from aggregating over high-frequency verbs, whereas the fast growing tail in the noun distributions appears to reflect the greater volume of low-frequency nouns.

In other words, the results imply that the differences observed between lexical categories do not necessarily warrant categorial distinctions. Rather, the observable differences appear to reflect the extent to which word co-occurrence clusters are shaped by the opposing communicative pressures of prediction and discrimination over the course of learning.

In other words, these results confirm the idea that lexical categories are not equally distributed across utterance positions. The next part of our analysis explores these relationships further.

#### *3.3. Lexical Category, Word Order, and Recurrence Patterns*

#### 3.3.1. What Makes a Lexical Category?

The distribution of function words suggests that function words will form the grammatical subcategory that is first discriminated systematically from the speech signal. As a consequence, it seems likely that, as both intuition and many linguistic theories would predict, function words provide a first contextual frame to aid in the learning of other grammatical and contextual categories. Once these basic contextual frames are learned, they will provide context, assisting in the learning of other words. The idea that context will provide information that aids learning in turn suggests lexical diversity will increase with utterance length.

Consistent with this suggestion, Genzel and Charniak [45] have shown that, although caching local probability estimates of a words' occurrence in written samples (to account for the variance in recurrence patterns over time) stabilizes relative entropy over lexical sample size in nouns significantly, the effect is far smaller in verbs and absent in function words. In the light of the foregoing discussion, this might be taken to suggest that patterns of co-occurrence in verbs are less variant than those in nouns and that these patterns are still less variant (and may even be regular) in function words. These considerations suggest in turn that the different subcategories of words systematically reduce uncertainty in communication at different levels of abstraction. To explore whether the different communicative contributions of words from different lexical categories are quantifiable in speech signal, we analyzed the patterns of occurrence of nouns, verbs, and function words (the three largest categories by token count) over utterance length.


*4.1.2 Operator/machine-related motion artifacts*

There are several operator/machine related artifacts that affect the accuracy and precision of OCTA measurements. Poorly focused images cause defocused artifacts (8A) that are caused by a decrease in reflective intensity resulting in reduced definition of retinal microvasculature Projection artifacts (8B) occur when the shadow of vessels in the superficial layer imprint onto the deeper layer. Segmentation errors (8C) are produced when the automated segmentation algorithm inaccurately identifies the layers of the retina or optic nerve. Decentration artifact (8D) occurs if the fovea or ONH is translocated to the periphery of the scan, preventing the algorithm from correctly identifying these areas. Cropping (8E) refers to loss in image data caused by B-scan not being fully within the capture window. Lastly, vitreous floaters may result in shadow artifacts (8F) because of decreased intensity of retinal layers in focused areas and may prevent clear visualization of structures in the affected area.

#### *4.1.3 Key factors that produce optical coherence tomography angiography artifacts*

Since OCTA artifacts may affect the accuracy of glaucoma diagnosis it is important to understand when and why these alterations are likely to occur. Older patients often have poor-quality scans due to a higher frequency of media opacities such as cataracts and age-related retinal pathologies. Recently Kamalipour et al. showed that 40% of OCTA images in the glaucoma group had poorer quality compared to healthy controls [47]. This was confirmed by Cheng et al. who found that the presence of glaucoma significantly increased the odds for the presence of any artifacts [48]. Similarly, patients with glaucoma were found to have two times the odds of incorrect automatic segmentation of the retinal superficial layer even after adjustment for age and VF loss [48]. Image quality was also found to decrease as glaucoma progressed. Declining retinal thickness reduces the signal intensity of reflected waves causing increased prevalence of artifacts.

#### *4.1.4 Prevalence of optical coherence tomography angiography artifacts*

In a well-diversified population of North Texas consisting of 292 patients, Mekala et al. found that most patients (99.3%) had at least one artifact [49]. The most common artifacts were due to eye movements (66.1%), defocus (64.7%) and shadows (40%). The most severe (affecting more than 10% of the scan area) were seen in patients with POAG (31.4%) and myopia (30.75%). A similar study by Weijing et al. found that 88.34% of images had at least one artifact, the most common of which was projection (100%), followed by motion (75.22%), blur (24.78%) and decentration (21.28%) [50]. Another study by Kamalipour et al. excluded 33.9% of scans due to poor image quality and found that 13.6% had one artifact and 9.8% had two or more. In this study the most common artifact was eye movement (10.6%), followed by defocus (9.6%), correctable segmentation errors (7.6%), and uncorrectable segmentation errors (5.4%) [48]. Given that multiple studies have confirmed the presence of artifacts in a significant percentage of images, it is crucial to account for artifacts when evaluating OCTA in a clinical setting.

#### *4.1.5 Current methods for reducing optical coherence tomography angiography artifacts*

Eye tracking technology was first introduced in 2004 as a way to compensate for eye movement and improve the quality of OCT scans [51]. Since then, this technology has shown to decrease the odds of acquiring poor-quality images by half [47]. It is highly effective in reducing eye movement and defocus artifacts but does not currently decrease the occurrence of segmentation artifacts.

Work by Venugopal et al. found that high-density (HD) scans, which increase sampling density from 304 × 304 A-scans to 400 × 400 A-scans, showed significantly greater VD values and fewer poor-quality scans than non-HD scans [52]. Thus, both eye tracking technology and HD scanning are proven methods for reducing the presence of OCTA artifacts. However, additional work is needed to further reduce the prevalence of other artifacts in OCTA.


**1. Introduction**

Celiac disease is defined as a lifelong condition as a result of ingestion of gluten among genetically susceptible people that can be relieved by the introduction of gluten-free diet; the condition relapses with gluten intake [1]. Recent studies show that it is prevalent around the world, covering from the western world, such as Europe and America, to Oceania, Africa, and Asia. The number of celiac sufferers increases, doubling its number every two decades [2]. The symptoms vary at a wide range, from flatulence, constipation, anorexia, irregular bowel habits, and irritability to numbness in limbs, foggy mind, diarrhea, and depression [3]. Therefore, it is hard to recognize the condition and deliver the diagnosis. In fact, almost 90% of celiac patients remain undiagnosed, due to the nonspecific or absent symptoms over a long period [4]. Thereby, it is of paramount significance to achieve the early-stage diagnosis of celiac disease that can lead to improving the patients' quality of life.

The current gold standard of celiac disease diagnosis, which has been also developed in a much earlier period, is to observe the small intestine atrophy obtained

through biopsy [1]. This process is highly invasive, time-consuming, and inconvenient to both patients and clinicians.

Over the last several decades, researchers have found that the concentration of some certain antibodies circulating in celiac patients' body increased, and the detection of celiac disease can be achieved with the detection of these antibodies. In fact, the surface of mucosa represents the major targeted sites when foreign antigens attack the body [5]. Plausibly, 80% of all cells producing immunoglobulin (IgG) in the human body are in small bowel mucosa, which also produces the dimers of IgA [6]. Therefore, the antibodies of celiac disease among untreated patients are located in the mucosal surface [7], as extracellular deposit, some present in jejunal juice [8], and most in the intestine [9, 10]. For untreated celiac patients, the additional generation of antibodies leads to an increase of antibodies specific to celiac disease (IgA− class), most of which can be found in the circulating blood and some other bodily fluids. Among all the celiac-specific antibodies, anti-reticulin (ARA), anti-jejunal (JEA), endomysial (EMA), and tissue transglutaminase antibodies (tTG) are among the patients' own endogenous biomolecules that form as a result of immune response to antigen in the intestine, whereas anti-gliadin antibody (AGA) and deamidated gliadin peptide antibody (DGP-Ab) are formed directly against dietary gliadin [11].

Over the years, the detection of the celiac-related antibodies has shown promising results, and whole blood- or serum-based pathology tests are regularly used for screening for celiac disease. Typically, screening for celiac disease includes tests for identification of titers for AGA and/or anti-tTG antibodies, and most of these tests are based on enzyme-linked immunoassays (ELISA). Even though the method of ELISA can reach a high sensitivity and specificity, these tests cannot be used on their own in the process of celiac disease diagnosis. Furthermore, pathology tests are typically confined to centralized laboratories, where expert personnel is required, leading to slow- and high-cost detections. Thereby, it is not suitable for the use outside hospitals, such as clinical offices or home settings, leaving a high number of undiagnosed celiac cases, especially when their symptoms are not obvious or do not affect their normal life. Thus, simple and rapid detection methods are in high demand to be developed.

Fast, accurate, and noninvasive early diagnosis methods and/or devices are needed to achieve the detection of celiac disease with high sensitivity and specificity, especially facing the rapid increasing number of celiac patients. Over the last decade, several point-of-care blood tests have been developed and applied for celiac diseases screening. The most prominent tests are Simtomax® Blood Drop system (Augurix SA, Switzerland) and Biocard™ celiac test (AniBiotech®, Finland), lateral flow immunoassays that detect anti-tTG and/or anti-deamidated gliadin peptide antibodies.

Lateral flow test, also called lateral flow immunoassay or test strip, has been widely and commercially used in the rapid detection of many diseases and conditions, such as HIV, illicit drugs, and early pregnancy [12]. In the following sections, we will discuss and compare several commercial kits available as POCT devices for celiac disease. The principle of lateral flow test is outlined in the next section.

## **2. Lateral flow immunoassays**

Lateral flow immunoassay is a simple immunochromatography technology that has successfully applied for rapid diagnostic testing [12, 13]. It typically made of nitrocellulose or paper-based porous membrane that makes it ideal to fabricate low-cost devices with little maintenance requirements. Porous membrane enables the separation, capture, and recognition of the target analytes. In addition, porous

**25**

**3. Point-of-care tests for celiac disease**

**3.1 Current commercial point-of-care tests**

*Challenges with Point-Of-Care Tests (POCT) for Celiac Disease*

*Schematic representation of lateral flow immunoassay in a capture format. Various important parts of the test are identified as sample, conjugate, and absorbent pads, nitrocellulose membrane with test and control lines.*

nature of the material facilitates movement of fluids, such as a whole blood, serum,

There are several types of lateral flow assays available, but the most popular is the capture format. There are two types of capture: "sandwich" and competitive

Overall, lateral flow immunoassay consists of four parts: sample pad, conjugate pad, nitrocellulose membrane (test line and control line labeled on it), and absorbent pad (also called wicking; **Figure 1**). Typically, fluid sample (blood, serum, urine, saliva) is added onto sample pad. Through the capillary action, the liquid moves to the conjugate pad, where preloaded recognition element (conjugated nanoparticles or colored reagent) is imbedded. The sample and the recognition reagent react commonly through antigen-antibody interaction. The sample together with the recognition element continues flowing within nitrocellulose membrane toward test and control lines. Depending on the type of a capture, sandwich or competitive, the test line would show a colorful line or disappear, respectively. Control line always shows colorful signal to indicate the correct functionality of the test. Absorbent pad function is to collect all the unreacted reagents as well as excess

Lateral flow immunoassay for celiac disease is typically used to detect antibodies, such as anti-tTG, anti-DGP, and AGA antibodies from the whole blood or serum in a sandwich type of detection [14–18]. Preloaded reagent can be gold nanoparticles or dye conjugated to antigens, such as transglutaminase, deamidated gliadin peptides, and gliadin protein fragments. The detection of celiac disease-related antibodies will present as color test line that can be usually seen by the naked eye. Lateral flow test or strip test is often used as a point-of-care test (POCT) due to its high specificity, visual color confirmation, and, especially, because of no additional instrumentation is required. In fact, most of the current commercial POCTs for celiac disease are based on lateral flow assay to detect antibodies. In the following section, we will discuss and make a comparison among commercial kits available together with the outlining principles of POCT device for celiac disease.

One of the most widely used commercial kits for celiac disease detection is the new generation of Biocard™ celiac test (AniBiotech®, Vantaa, Finland). In

or urine, by means of capillary action with no external force required.

*DOI: http://dx.doi.org/10.5772/intechopen.81874*

**Figure 1.**

capture [13].

of liquids.

*Challenges with Point-Of-Care Tests (POCT) for Celiac Disease DOI: http://dx.doi.org/10.5772/intechopen.81874*

**Figure 1.**

*Celiac Disease - From the Bench to the Clinic*

nient to both patients and clinicians.

are formed directly against dietary gliadin [11].

in high demand to be developed.

**2. Lateral flow immunoassays**

through biopsy [1]. This process is highly invasive, time-consuming, and inconve-

Over the last several decades, researchers have found that the concentration of some certain antibodies circulating in celiac patients' body increased, and the detection of celiac disease can be achieved with the detection of these antibodies. In fact, the surface of mucosa represents the major targeted sites when foreign antigens attack the body [5]. Plausibly, 80% of all cells producing immunoglobulin (IgG) in the human body are in small bowel mucosa, which also produces the dimers of IgA [6]. Therefore, the antibodies of celiac disease among untreated patients are located in the mucosal surface [7], as extracellular deposit, some present in jejunal juice [8], and most in the intestine [9, 10]. For untreated celiac patients, the additional generation of antibodies leads to an increase of antibodies specific to celiac disease (IgA− class), most of which can be found in the circulating blood and some other bodily fluids. Among all the celiac-specific antibodies, anti-reticulin (ARA), anti-jejunal (JEA), endomysial (EMA), and tissue transglutaminase antibodies (tTG) are among the patients' own endogenous biomolecules that form as a result of immune response to antigen in the intestine, whereas anti-gliadin antibody (AGA) and deamidated gliadin peptide antibody (DGP-Ab)

Over the years, the detection of the celiac-related antibodies has shown promising results, and whole blood- or serum-based pathology tests are regularly used for screening for celiac disease. Typically, screening for celiac disease includes tests for identification of titers for AGA and/or anti-tTG antibodies, and most of these tests are based on enzyme-linked immunoassays (ELISA). Even though the method of ELISA can reach a high sensitivity and specificity, these tests cannot be used on their own in the process of celiac disease diagnosis. Furthermore, pathology tests are typically confined to centralized laboratories, where expert personnel is required, leading to slow- and high-cost detections. Thereby, it is not suitable for the use outside hospitals, such as clinical offices or home settings, leaving a high number of undiagnosed celiac cases, especially when their symptoms are not obvious or do not affect their normal life. Thus, simple and rapid detection methods are

Fast, accurate, and noninvasive early diagnosis methods and/or devices are needed to achieve the detection of celiac disease with high sensitivity and specificity, especially facing the rapid increasing number of celiac patients. Over the last decade, several point-of-care blood tests have been developed and applied for celiac diseases screening. The most prominent tests are Simtomax® Blood Drop system (Augurix SA, Switzerland) and Biocard™ celiac test (AniBiotech®, Finland), lateral flow immunoassays that detect anti-tTG and/or anti-deamidated gliadin peptide antibodies. Lateral flow test, also called lateral flow immunoassay or test strip, has been widely and commercially used in the rapid detection of many diseases and conditions, such as HIV, illicit drugs, and early pregnancy [12]. In the following sections, we will discuss and compare several commercial kits available as POCT devices for celiac disease. The principle of lateral flow test is outlined in the next section.

Lateral flow immunoassay is a simple immunochromatography technology that has successfully applied for rapid diagnostic testing [12, 13]. It typically made of nitrocellulose or paper-based porous membrane that makes it ideal to fabricate low-cost devices with little maintenance requirements. Porous membrane enables the separation, capture, and recognition of the target analytes. In addition, porous

**24**

*Schematic representation of lateral flow immunoassay in a capture format. Various important parts of the test are identified as sample, conjugate, and absorbent pads, nitrocellulose membrane with test and control lines.*

nature of the material facilitates movement of fluids, such as a whole blood, serum, or urine, by means of capillary action with no external force required.

There are several types of lateral flow assays available, but the most popular is the capture format. There are two types of capture: "sandwich" and competitive capture [13].

Overall, lateral flow immunoassay consists of four parts: sample pad, conjugate pad, nitrocellulose membrane (test line and control line labeled on it), and absorbent pad (also called wicking; **Figure 1**). Typically, fluid sample (blood, serum, urine, saliva) is added onto sample pad. Through the capillary action, the liquid moves to the conjugate pad, where preloaded recognition element (conjugated nanoparticles or colored reagent) is imbedded. The sample and the recognition reagent react commonly through antigen-antibody interaction. The sample together with the recognition element continues flowing within nitrocellulose membrane toward test and control lines. Depending on the type of a capture, sandwich or competitive, the test line would show a colorful line or disappear, respectively. Control line always shows colorful signal to indicate the correct functionality of the test. Absorbent pad function is to collect all the unreacted reagents as well as excess of liquids.

Lateral flow immunoassay for celiac disease is typically used to detect antibodies, such as anti-tTG, anti-DGP, and AGA antibodies from the whole blood or serum in a sandwich type of detection [14–18]. Preloaded reagent can be gold nanoparticles or dye conjugated to antigens, such as transglutaminase, deamidated gliadin peptides, and gliadin protein fragments. The detection of celiac disease-related antibodies will present as color test line that can be usually seen by the naked eye.

Lateral flow test or strip test is often used as a point-of-care test (POCT) due to its high specificity, visual color confirmation, and, especially, because of no additional instrumentation is required. In fact, most of the current commercial POCTs for celiac disease are based on lateral flow assay to detect antibodies. In the following section, we will discuss and make a comparison among commercial kits available together with the outlining principles of POCT device for celiac disease.