**4. Conclusions**

If the characteristics of the people are added together with the possibilities that the environment implies, it can generate or promote a level of activity over the older

#### *Not All Rehabilitation is Physical Therapy DOI: http://dx.doi.org/10.5772/intechopen.94599*

adult, allowing to give sense to variables or psychological aspects in an intrinsic or extrinsic way in a positive or negative way against a stimulus or activity adapted to their characteristics [33].

Factors such as psychological well-being as a way of accepting oneself and developing one's own potential for personal growth and capacity to face life events, together with self-efficacy as a way of motivating performance, can increase both physical and psychological possibilities in the field of geriatrics.

Finally, as the title of the chapter shows, not everything is physical rehabilitation, since other factors associated with the person must be taken into account, which can play an important role, not only in the predisposition towards a treatment or intervention, but also in the normal development of his/her day-to-day life.


Summary

In summary, CS-derived drug-loaded nanomaterial has been shown to have a reasonable encapsulation efficiency, an appropriate hydrodynamic diameter, manageable surface charge, low toxicity, and improved anticancer properties compared to the free drug [149,153,154].


**5.1 Methods applied in continental slope of the Campos Basin (SE, Brazil)**

In the Oceanprof project, 41 surface sediment samples were collected by using a box corer, along transects ranging from 750 to 1950 m of water depth on the continental slope (**Figure 5**), in the austral winter of 2003. The core top (0–2 cm interval) sediment sampled in each site was used to understand the living and dead benthic foraminifera distribution patterns and ecological preferences. For this purpose, identification and quantitative foraminifera analysis were performed using the 63 μm size fraction. The variables considered for the statistical analysis included percentages of sand and mud, calcium carbonate and total organic carbon contents in the sediment, and total phosphate in the water (see details in [73]).

In the same project (Oceanprof), 20 surface sediment samples were chosen along 1050 and 1950 m depth (**Figure 5**). In this case, only living (rose Bengal stained) benthic foraminifera were analyzed in combined samples collected in four different slices of the cores (0–2, 2–5, 5–10, and 10–15 cm) to understand the distribution patterns and their ecology. In this study, the variables considered for statistical analysis included the sand and mud contents, particulate organic matter flux to the sea floor, bottom water dissolved oxygen concentrations, calcium carbonate, total organic carbon, total nitrogen, and total lipid contents in the sediment (see details in [75]).

 In the Habitats project, an ecological study of living (rose Bengal stained) benthic foraminifera was performed in samples collected with a box corer, on the continental slope, Plateau of São Paulo, and canyons, during two campaigns (austral winter of 2008 and summer of 2009). The stations followed nine transects from 400 to 3000 m deep (**Figure 5**). In the Grussaí and Almirante Câmara canyons, the stations were located in four isobaths (400, 700, 1000, and 1300 m deep) and the obtained data were compared with adjacent transects on the open slope (**Figure 5**). Changes in the density, diversity, and composition of benthic foraminifera were analyzed in response to environmental factors (i.e., sand and mud contents, calcium carbonate, total organic carbon and chlorophyll-*a* concentrations, and phytopigment concentrations in the sediment) (see details in [76]).

### **5.2 Results obtained in continental slope of the Campos Basin and discussion**

 The middle slope is characterized by the dominance of different species of the genus *Bolivina, Cassidulina laevigata*, and *Globocassidulina subglobosa*. The occurrence of these species in association with *Cibicidoides kullenbergi, Epistominella exigua*, and *Uvigerina proboscidea* seems to be related to seasonal organic matter fluxes, relatively oxic bottom waters, strong bottom currents, and sandy sediments. The lower slope is inhabited by a microfauna with different characteristics, preferentially composed of epifaunal or shallow infaunal deposit feeding species (e.g., *Bolivina* spp., *Eponides weddellensis,* and *Lenticulina cultrata*) and suspension feeders that are adapted to oligotrophic conditions and high dissolved oxygen levels in the bottom waters, for example, *Rhabdammina* spp., *Rhizammina* sp. [73].

Yamashita et al. [75] concluded that besides the sediment grain size, the vertical flux of particulate organic matter seems to be the main factor controlling the spatial distribution of benthic foraminifera species in the slope of Campos Basin. The middle slope (1050 m of water depth) was characterized by relatively high

foraminiferal density and a predominance of phytodetritus-feeding foraminifera such as *Epistominella exigua* and *Globocassidulina subglobosa*. The occurrence of these species seems to reflect the Brazil Current System (BCS). The abovementioned currents are associated with the relatively high vertical flux of particulate organic matter and the prevalence of sandy sediments, respectively. The lower slope (between 1350 and 1950 m of water depth) was marked by low foraminiferal density and assemblages composed of *Bolivina* spp. and *Brizalina* spp., with low particulate organic matter flux values, muddy sediments, and more refractory organic matter. The distribution of this group seems to be related to episodic fluxes of food particles to the seafloor, which are influenced by the BCS at the surface and are deposited under low deep current activity (Intermediate Western Boundary Current; [75]).

According to Sousa et al. [76], the availability and quality of the food, the energy state (stability) at the benthic/pelagic boundary, and the grain size of the substrate seem to be the most important environmental factors determining the distribution pattern of the benthic foraminiferal assemblages in the Campos Basin slope. The highest values of density, diversity, and richness, as well as the predominance of hyaline calcareous foraminifera and infaunal species, reflect a higher contribution of food received continuously in the shallower stations (400 m depth). At 700–1000 m of water depth, the density of foraminifera decreased and there was a larger presence of opportunistic species, possibly reflecting the pulse of phytodetritus. The considerable increase in the agglutinated foraminifera, the continuous decrease in the density of foraminifera, and also decrease in the values of the Benthic Foraminifera High Productivity Index (BFHP [61]) as depth increases indicate typical oligotrophic conditions in this sector of the Campos Basin (lower slope and São Paulo Plateau).

The comparison of density and species composition data in the austral winter of 2008 and austral summer of 2009 periods allows us to infer that during the winter, the food input was higher. The values of density and biomass of living benthic foraminifera allow us to suggest that the Almirante Câmara canyon is a greater entrapment site of organic matter between 400 and 1000 m isobaths in comparison to the open areas at the same isobaths [76].

Even though the stations analyzed were the same or spatially very close to each other, the response of benthic foraminifera of the Campos Basin seems to be slightly different due to the seasonality. Because of these dissimilar times, the local hydrodynamics can change and, consequently, variations can occur in the particulate organic matter flux, and quantity and quality of the organic matter on the sea floor [73, 75, 76]. The benthic foraminifera and the geochemical results of these projects showed that, despite the intense oil exploration and production activities in the Campos Basin by PETROBRAS, these specific study areas (**Figure 5**) did not suffer anthropic impact in terms of pollutants [77].


*2.2. Pre-clinical AAV-mediated Gene Therapy for PFIC3*

ABCB4 deficiency seems the best option to investigate the feasibility of gene therapy as a treatment option for PFIC. This is because of the availability of a suitable animal model and a previous study showing that partial correction by hepatocyte transplantation provides therapeutic correction [52]. The *Mdr*2 −/− (*Abcb4*−/−) mouse has been demonstrated to be a relevant model for PFIC3 [10,53]. The severity of the pathology depends on the mouse strain, with FVB mice displaying a more severe phenotype compared to the C57Bl/6 mice. Adjustments via the diet, such as cholate supplementation, increase the mouse bile hydrophobicity which worsens the disease phenotype [54]. In a recent study we investigated the feasibility of AAV8-h*ABCB4* mediated correction in adult *Abcb*4 −/− mice with a C57Bl/6 background [55]. Upon vector administration, the efficacy was monitored over time until 6 months using dietary cholate administration to mimic the human bile toxicity [54]. This study provided proof of concept by demonstrating long-term correction, as shown by normalization of the liver damage parameters AST, Alanine Transaminase (ALT) and Alkaline Phosphatase (ALP), and the absence of fibrosis. Restoring a sufficient Phosphatidylcholine (PC) content in bile is a pre-requisite for prolonged correction using AAV. In case of insufficient correction, the ongoing hepatocyte proliferation will result in loss of AAV vectors, thereby further reducing PC presence in bile and thus increasing the liver damage. This was demonstrated by Weber et al. with *Abcb4*−/− FVB mice displaying a more severe phenotype [56]. These mice were treated with an AAV vector, consisting of a codon-optimized h*ABCB4* at week 2 after birth and the effect was monitored for 12 weeks. In males, this treatment appeared effective, but in 50% of the females the correction was lost. To overcome this, a second cohort was injected two times, first at week 2 and three weeks later a second dose was given. This protocol provided prolonged correction and is, with regard to severity of the pathological model, more comparable to PFIC3 patients. Both studies do indicate that long-term correction of a disease causing hepatocyte proliferation using non-integrating AAV vectors is feasible but only if the efficacy is sufficient.

In PFIC3 patients, the disease onset is observed in young children and therefore ideally the therapy should be given at an early stage. This complicates the use of a nonintegrating vector like AAV. Several studies have shown that the initial correction is lost over time when treating neonatal or juvenile animals [49,50]. Moreover, the induction of proliferation upon (partial) loss of correction will contribute to episomal transgene loss and will accelerate the decrease in therapeutic efficacy. This hurdle can be overcome by strategies that aim for integration of the therapeutic transgene. Siew et al. tested integration of a therapeutic construct, consisting of a liver specific promoter and a codonoptimized human *ABCB4* transgene, flanked by the piggyBac transposase short terminal repeats [57]. Co-administration of this construct with an AAV2/8 vector containing a piggybac expression cassette to juvenile FVB *Abcb*4 −/− mice resulted in the integration of the therapeutic construct providing lifelong hABCB4 expression and correction of

the disease. Integration is non-random, but transposons do cause integration in close proximity of actively transcribed gene regions, transcription start sites, and open chromatin structures [58,59]. None of the analyzed unique integration regions were linked to genes known to play a role in hepatocellular carcinomas [59]. Nevertheless, this cannot be excluded. Targeting the transgene integration to a safe genomic locus could overcome this safety issue.

An obvious target to ensure integration in an active region in hepatocytes is the Albumin locus. A recent study aimed at Homologous Directed Repair (HDR) mediated integration of a promoterless Factor IX coding region in this locus without using a nuclease [60]. As the episomal construct lacks a promoter, and a self-cleaving protein is added to the transgene, its expression is regulated by the expression of albumin. The absence of a promoter and a nuclease increases safety of this "generide" strategy. The basis of this approach is to deliver a Factor IX encoding transgene flanked by two homologous arms to the albumin gene with complimentary sequences coding for the desired integration region. Using the generide strategy Muro et al. demonstrated that this could be used to correct a metabolic disorder, albeit with a low efficacy [61]. Inducing double strand breaks in this locus will stimulate HDR and integration efficacy. In a follow up study, increased correction was established by combining the generide approach with an AAV-*saCas9* guided to the albumin locus [62]. In all studies neonatal mice were used, modelling the use early after birth, when the liver is actively growing and hepatocytes are proliferating. The latter is a major advantage because during cell division the HDR system is active whereas it is inactive in quiescent cells [63]. In this study, life-long therapeutic correction of the pathology was established with a treatment shortly after birth. Further, no off-target integrations were seen in predicted sites which underlines the safety profile of this gene modulation system [62]. This targeted integration strategy seems a feasible option to treat PFIC shortly after birth.


**7. Conclusions**

A novel method for solving the very important "matrix inversion" problem has been developed and validated in this paper. Consequently, it can also solve linear algebraic systems of equations. The concept developed is essentially an RNN-based analog computing machine.

We have compared this novel method with other dynamical systems methods from the relevant literature. It has been demonstrated that our novel method does theoretically have an exponential convergence rate towards the exact solution of the problem for any IVP (initial value problem). Also, the convergence rate of this method is much higher than those of other related competing concepts.

It is further possible to customize this novel model in order to enable its implementation on a cellular neural network processor machine. This has previously been done in our previous works.

For validation we have extensively compared our method with other relevant competing methods like gradient descent, Zhang, and Chen methods in one large simulation experiment. Hereby, we have considered the following scenarios: 1000 random matrixes of M are generated and applied in the different dynamical system models for matrix inversion. For each time-point, errors reached are summed-up using the formula *MX* − *I*.

It has been clear that while using our novel method we do visibly reach a significant speed-up even on one single CPU, this compared to the other methods. A use of more coefficients (i.e., higher values for n in model Equations (24) and (31)) will surely result in an even much higher convergence rate. On the other hand, however, if we use more coefficients, we would need more preparation time to create the templates and consume more computing resources for running the (our novel) RNN processor dynamical model. Therefore, it is recommended to reach a balance/tradeoff between convergence rate and number of required coefficients (i.e., value of n).

To finish, the last experiments have demonstrated that using the polynomial function for F in Equation (31) does lead to a clearly much higher convergence rate when compared to the other types of function. Further, the higher the polynomial order, the best.

**Author Contributions:** Conceptualization, V.T. and K.K.; Methodology, J.C.C. and K.K.; Software, V.T.; Validation, V.T., J.C.C. and K.K.; Formal Analysis, V.T.; Investigation, V.T.; Resources, V.T.; Data Curation, V.T.; Writing-Original Draft Preparation, V.T.; Writing-Review & Editing, J.C.C. and K.K; Visualization, V.T.; Supervision, J.C.C. and K.K.; Project Administration, K.K.

**Funding:** This research received no external funding.

**Conflicts of Interest:** The authors declare no conflict of interest