Cell transplantation therapy is a potentially powerful tool and can be used to replace defective cells with healthy cells. This offers the possibility of alleviating the destructive symptoms for many diseases such as Parkinson's disease, Alzheimer's disease, stroke, spinal cord trauma, Type I diabetes and many more. While there are many diseases that could be positively impacted from cell transplantation therapy, the focus of this research is insulin dependent, Type I Diabetes. The Islets of Langerhans are composed of various types of cells located in the pancreas and are responsible for a variety of biochemical functions. Specifically, the beta Islet cells are responsible for production of the hormone insulin that regulates and aids in biosynthesis of glucose. Transplantation of isolated allografted pancreatic islets, which contain insulin producing cells, into diabetic rats has proven to be highly successful. However, these transplantations involve using medications for long term immunosuppression to defend against an undesired host immune response. Immunosuppressive medications are both costly and illicit additional side effects that can be detrimental to the host. This research focuses on the use of testicular derived Sertoli cells that have been publicized to provide localized immunoprotection. Electrofusion is a process that can be used to fuse homogeneous and heterogeneous cell types by promoting the creation of micropores in the cell's lipid bilayer. This renders the cell temporarily fusogenic, or capable of facilitating fusion. Cells must then be brought into contact with one another via mechanical, chemical or viral means. This research study proposes to optimize electrofusion technology to create novel, secretory hybrids composed of Islet and Sertoli cells that are immunoprotected and produce insulin in response to a glucose challenge. The components of the electrofusion device include a Sterlitech 0.2 ìm microporous membrane, a woven cellulose absorbent pad, two aluminum electrodes and a chamber body and top injection molded using Delrin. Preliminary experiments using B16-F10 murine melanoma cells incorporated with centrifugation to increase cell to cell contact resulted in an average fusion yield of 18.9% ± 8.1 SD using a field strength of 2500 V/cm, 8 pulses and a 250 ìs pulse length. Additionally, lab synthesized electroporation buffers containing 8.5% sucrose (w/v) and 0.3% glucose increased total and viable fusion yields to 37.1% ± 9.3 SD and 13.8% ± 2.1 SD, respectively. These results showed promise and should be further validated with additional cell lines and tissues to corroborate reproducibility.

No description available.

The course of microsporidiosis caused by Encephalitozoon cuniculi in immunocompetent BALB/c mice and immunodeficient SCID mice was screened using molecular methods. The site of infection in organs was located using molecular and histology methods. The effectiveness of albendazole treatement and possibility of infection relapse after immunosuppresion (cyclosporine A, tacrolimus, mycofenolate mofetil) was also studied. Moreover, the course of excretion of microsporidial spores in feces was monitored during the whole time of experiment.

Indiana University-Purdue University Indianapolis (IUPUI) / Sepsis is a clinical syndrome due to a systemic inflammatory response to severe microbial infection. Little is known about the changes in the bone marrow (BM) and how they affect the hematopoietic response to bacterial infection. Using an animal model of severe sepsis induced by Pseudomonas aeruginosa, we have previously reported that hematopoietic stem cells (HSC) undergo a significant expansion in the BM accompanied with myeloid suppression. This bone marrow response was Toll-like Receptor 4 (TLR4)-dependent. TLR4 is activated by bacterial lipopolysaccharide (LPS) and signals through two major independent downstream molecules: TRIF and MyD88. In the present study, I found that the TLR4/TRIF and the TLR4/MyD88 pathways contribute in a distinct manner to the BM response to P. aeruginosa's LPS. TRIF plays a major role in the expansion of the HSC pool, whereas MyD88 is required for myeloid suppression. Following LPS stimulation, HSCs enter in the cell cycle, expand and exhaust when transplanted in healthy mice. Loss of TRIF rescued completely the long-term engraftment and multilineage reconstitution potential of septic HSCs, but did not affect myeloid differentiation. Conversely, MyD88 deficiency prevented completely the myeloid suppression in the myeloid progenitors, but conferred limited protective effects on the HSC function. It is of great therapeutic value to identify the downstream molecules involved in TLR4/MyD88 dependent myeloid suppression. I found miR-21, a microRNA that is involved in inflammation, was up-regulated upon LPS challenge in a MyD88-dependent manner. However, deletion of miR-21 in the BM did not rescue LPS-induced bone marrow dysfunction, demonstrating that miR-21 is not a critical regulator in these processes. Further studies are warranted to determine the precise molecular mechanisms involved in the complex pathogenesis of BM response to sepsis. Taken together, my results show for the first time that the TLR4/TRIF signaling as a key mediator of HSC damage during acute LPS exposure and that activation of the TLR4/MyD88 signaling pathway play a dominant role in myeloid suppression. These results provide novel insights into our understanding of the molecular mechanisms underlying bone marrow injury during severe sepsis and may lead to the development of new therapeutic approaches in this disease.