Publications (10)14.69 Total impact
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Article: In-vitro generation of human adipose tissue derived insulin secreting cells: up-regulation of Pax-6, Ipf-1 and Isl-1.
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ABSTRACT: We present a study of up-regulation of genes responsible for pancreatic development in glucose-sensitive insulin-secreting mesenchymal stem cells (IS-MSC) generated and differentiated from human adipose tissue (h-AD), with use of our specific differentiation media and without use of any xenogenic material. Anterior wall abdominal fat was collected from 56 volunteers and cultured in self-designed proliferation medium for 10 days. Cells were harvested by trypsinization and differentiated into insulin-expressing cells using self-designed differentiation medium for 3 days followed by evaluation for transcriptional factors Pax-6, Ipf-1, Isl-1, C-peptide and insulin secretion. Generated IS-MSC showed expression of Pax-6, Pdx-6 and Isl-1. Non-differentiated MSC as well as their further culture in absence of differentiation medium were used as negative controls. Generated 56 IS-MSC cell-lines were glucose responsive i.e. mean C-Peptide and insulin secretion levels were measured 0.41 ng/ml and 13.13 μU/ml, respectively, in absence of glucose which rose to 1.18 ng/ml and 83.42 μU/ml, respectively, following glucose challenge (p < 0.001). The mean rise in C-peptide and insulin secretion levels was 2.88 and 6.35 fold, respectively. To conclude insulin-secreting h-AD-MSC can be generated safely and effectively with application of specific differentiation media without xenogeneic material/any genetic modification, showing expression of transcriptional factors Pax-6, Ipf-1 and Isl-1.Cytotechnology 05/2013; · 1.21 Impact Factor -
Article: Stem Cells Versus Donor Specific Transfusions for Tolerance Induction in Living Donor Renal Transplantation: A Single-Center Experience.
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ABSTRACT: BACKGROUND: We undertook this study to define the role of stem cell transplantation (SCT) versus donor-specific transfusion (DST) in tolerance induction and sustenance in living donor renal transplantation (LDRT). METHODS: In this prospective three-armed trial in LDRT with 13 patients each in demographically well-balanced groups, tolerance induction protocol (TIP) was used with SCT in group 1, DST in group 2, and no induction in group 3. Tolerance induction protocol consisted of SCT/DST under conditioning with bortezomib, methylprednisone, rituximab, and rabbit antithymocyte globulin. Transplantation was performed with prednisone in groups 1 and 2 and with triple immunosuppression in group 3, if lymphocyte/flow crossmatch was negative; and if donor-specific antibodies (DSAs) were absent in the first 2 groups. Posttransplant monitoring included serum creatinine (SCr), peripheral T-regulatory cells (pTregs)(127/CD4+/25), and DSA for groups 1 and 2; DSA was eliminated in group 3. Rescue IS was started with rise of SCr/DSA/ rejection. RESULTS: Tolerance induction protocol was safe. Over a mean follow-up of 2 years, no patient/graft was lost in groups 1 and 2. One patient of group 3 lost graft to noncompliance. Protocol biopsies were unremarkable. Rejections were noted in six patients of group 1, five of group 2, and seven of group 3. Donor-specific antibodies were elevated in three patients of both groups. Mean SCr of all groups was similar; however, pTregs were increased posttransplant in groups 1 and 2 versus group 3. Group 1 had sustained rise in pTregs. CONCLUSION: Stem cell transplantation and DST are useful for immunosuppression minimization in LDRT with sustained generation of pTregs with SCT.Transplantation 01/2013; 95(1):155-160. · 4.00 Impact Factor -
Article: Single-center experience on renal transplantation in primary focal and segmental glomerulosclerosis using hematopoietic stem cell transplantation in thymus, bone marrow, portal and peripheral circulation.
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ABSTRACT: Recurrence of primary focal segmental glomerulosclerosis (FSGS) is an important cause of graft loss after renal transplantation (RTx). We report our experience in 34 patients with primary FSGS who underwent RTx between April 1999 and June 2009, using hematopoietic stem cell transplantation (HSCT). They belonged to four groups: group 1 (n = 12) received high-dose HSCT in periphery, thymus, bone-marrow, and portal circulation with low-dose non-myeloablative conditio-ning; group 2 (n = 7) was modified with HSCT without marrow/thymic infusion; and group 3 (n = 3) received HSCT and proteasome inhibitor Bortezomib replacing conditioning. Group 4 (n = 12), were controls who opted for RTx under standard triple-drug immunosuppression. Patient/donor demogra-phics were comparable in all. No recurrence was noted in group 1 with mean follow-up of 8.1 years, whereas 28.6% of group 2, 33.3% of group 3, and 36.4% of group 4 had recurrence over mean follow-up of 2.6, 1.1, and 6.5 years, respectively. Mean serum creatinine was 1.62, 1.69, 1.41, and 1.73 mg%, respectively. Rejections were noted in 41.7%, 28.6%, 0%, and 45.5% grafts, respectively. Groups 1 and 4 had 25% patient loss each, group 2 had 28.6% loss, and no loss was observed in group 3. Graft loss was noted in 33.3% in group 1, 14.3% in group 2, nil in group 3, and 16.7% in the last group. Recurrent FSGS was prevented in RTx with HSCT in thymic, marrow infusion under low-dose non-myeloablative conditioning compared to controls and Bortezomib group, thus suggesting potential role of central tolerance in FSGS.Saudi journal of kidney diseases and transplantation: an official publication of the Saudi Center for Organ Transplantation, Saudi Arabia 01/2013; 24(1):15-21. -
Article: Co-infusion of adipose tissue derived mesenchymal stem cell-differentiated insulin-making cells and haematopoietic cells with renal transplantation: a novel therapy for type 1 diabetes mellitus with end-stage renal disease.
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ABSTRACT: Type 1 diabetes mellitus (T1DM) is a common cause of end-stage renal disease (ESRD). Various factors contribute to wide fluctuations in blood glucose levels and exogenous insulin requirement in such patients even after renal transplantation (RT). Simultaneous pancreas-kidney transplantation is one of the therapies for these patients. Stem cell (SC) therapy for T1DM and for minimisation of immunosuppression after RT has shown encouraging results. We report a 30-year-old-man with T1DM since 15 years and ESRD since 2 years, who underwent living donor RT and co-infusion of in vitro generated insulin-making cells differentiated from donor adipose tissue derived mesenchymal stem cells and bone marrow -derived haematopoietic SC into subcutaneous tissue, portal and thymic circulation under non-myeloablative conditioning. Over follow-up of 13 months he has stable graft function with serum creatinine, 1.2 mg/dl, zero rejection and glycosylated haemoglobin level of 6.1% on calcineurin-inhibitor based therapy.Case Reports 01/2013; 2013(may23_1). -
Article: Stable graft function on low-dose steroid monotherapy in spite of donor-specific antibodies in renal transplantation combined with stem cell infusion.
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ABSTRACT: Transplantation using immunosuppression/induction therapies has controlled acute rejections; however, there is no answer for chronic graft attrition. Donor-specific antibodies (DSA) are believed to cause antibody mediated rejections eventually causing chronic graft loss. Regulatory T cells (T-regs) are believed to protect the graft from immune injury. We report a 53-year-old woman transplanted with her son's kidney using donor-specific transfusion and stem cells (SC) under non-myeloablative conditioning of cyclophosphamide, anti-T and anti-B-cell antibodies and Bortezomib. The patient was on low-dose steroid monotherapy under immune monitoring of DSA and serum creatine. Graft biopsy at 1 and 3.5 years post-transplant was unremarkable in spite of the presence of DSA. Peripheral T-regs (pTregs) at 3.5 years post-transplant were 3.54%. This case shows that DSA are not necessarily detrimental to the renal allograft. We further hypothesise that pTregs were induced from SC and sustained to protect this graft from cytotoxic T cells and DSA.Case Reports 01/2013; 2013. -
Article: Ex vivo generation of glucose sensitive insulin secreting mesenchymal stem cells derived from human adipose tissue.
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ABSTRACT: Diabetics are incapable of producing insulin/have autoimmune mechanisms making it ineffective to control glucose secretion. We present a prospective study of glucose-sensitive insulin-secreting mesenchymal stem cells (IS-MSC) generated from human adipose tissue (h-AD) sans xenogenic material. Ten grams h-AD from donor anterior abdominal wall was collected in proliferation medium composed of α-Minimum Essential Media (α-MEM), albumin, fibroblast-growth factor and antibiotics, minced, incubated in collagenase-I at 37°C with shaker and centrifuged. Supernatant and pellets were separately cultured in proliferation medium on cell+ plates at 37°C with 5% CO(2) for 10 days. Cells were harvested by trypsinization, checked for viability, sterility, counts, flow-cytometry (CD45(-)/90(+)/73(+)), and differentiated into insulin-expressing cells using medium composed of DMEM, gene expressing up-regulators and antibiotics for 3 days. They were studied for transcriptional factors Pax-6, Isl-1, pdx-1 (immunofluorescence). C-peptide and insulin were measured by chemiluminescence. In vitro glucose sensitivity assay was carried out by measuring levels of insulin and C-peptide secretion in absence of glucose followed by 2 hours incubation after glucose addition. Mean IS-AD-MSC quantum was 3.21 ml, cell count, 1.5 ×10(3) cells/μl), CD45(-)/90(+)/73(+) cells were 44.37% /25.52%. All of them showed presence of pax-6, pdx-1, and Isl-1. Mean C-Peptide and insulin levels were 0.36 ng/ml and 234 μU/ml, respectively, pre-glucose and 0.87 ng/ml and 618.3 μU/ml post-glucose additions. The mean rise in secretion levels was 2.42 and 2.65 fold, respectively. Insulin-secreting h-AD-MSC can be generated safely and effectively showing in vitro glucose responsive alteration in insulin and C-peptide secretion levels.Indian journal of endocrinology and metabolism. 03/2012; 16 Suppl 1:S65-9. -
Article: Effect of co-transplantation of mesenchymal stem cells and hematopoietic stem cells as compared to hematopoietic stem cell transplantation alone in renal transplantation to achieve donor hypo-responsiveness.
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ABSTRACT: We evaluated donor hypo-responsiveness in renal allograft recipients to donor adipose tissue-derived mesenchymal stem cell (h-AD-MSC) +hematopoietic stem cell transplantation (HSCT) vs. HSCT alone. Patients were divided into 2 demographically equal groups (n = 100) A and B subjected to equal non-myeloablative conditioning of target-specific irradiation, anti-T + B cell antibodies and cyclophosphamide with HSCT. Group A was administered h-AD-MSC additionally. Transplantation was performed following favorable cross-matching. Cyclosporine, 3 mg/kg BW/day + prednisone, 20 mg/day were immunosuppressants for first 3 months, cyclosporine was replaced by azathioprine subsequently and prednisone lowered to 5-10 mg/day. Peripheral blood chimerism (PBC) was studied using fluorescent in situ hybridization technique at 3/18 months post transplant. Biopsy was performed for graft dysfunction and reported as per Banff criteria,'05. Mean nucleated HSC counts (n × 10(8)/kgBW) was 7.32 with mean CD34+ yield 0.09% in group A; and 6.98 and 0.40% in group B, respectively; CD45-/90+ was 13.49% in former. Over 18 months post transplant, former had mean serum creatinine (SCr), 1.59 mg%, 12% acute rejection (AR) episodes, 3% patient, 1% patient +graft loss; latter had mean SCr 1.49 mg%, 18% AR episodes, 1% patient, 6% graft and 8% patient +graft losses. PBC was higher (4%) in former than later (1.8%). Combined h-AD-MSC +HSCT under non-myeloablative conditioning was safe, more effective than HSCT alone to achieve donor hypo-responsiveness with adequate stable graft function and reduced rejection episodes.International Urology and Nephrology 03/2011; 43(1):225-32. · 1.47 Impact Factor -
Article: Treatment of polyglandular autoimmune syndrome type 3 using co-transplantation of insulin-secreting mesenchymal stem cells and haematopoietic stem cells.
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ABSTRACT: The authors report a 17-year-female and a 19-year-male with uncontrolled insulin-dependent diabetes mellitus (IDDM) for ≥10 years, treated with insulin-secreting human adipose tissue derived mesenchymal stem cells (IS-h-ADMSC). Both had hypothyroidism and were diagnosed as polyglandular autoimmune syndrome type-3 (PGAS-3). PGAS are rare polyendocrinopathies with ≥2 endocrine disorders mediated by autoimmune mechanisms leading to hypo-function and organ failure. Therapeutic options are hormone replacement, immunosuppression and avoiding infection. The authors administered autologous H-AD-IS-MSC+bone marrow-derived haematopoietic stem cells (HSC) into portal circulation with conditioning of cyclophosphamide, bortezomib, rituximab and rabbit-antithymoglobulin. Over follow-up of 38 and 16 months, respectively, both are doing well with sustained fall of glycosylated haemoglobin (Hb1Ac) from 8.1 to 6.4% and 14.2 to 8.6%, respectively and C-peptide raised from 0.01 to 0.23 ng/ml and 0.1 to 0.34 ng/ml, respectively with sustained 40% decreased insulin requirement. Thus long-term control of IDDM in PGAS-3 with co-transplantation of H-AD-IS-MSC+HSC can be achieved safely and effectively.Case Reports 01/2011; 2011. -
Article: Clonal deletion with bortezomib followed by low or no maintenance immunosuppression in renal allograft recipients.
Transplantation 07/2010; 90(2):221-2. · 4.00 Impact Factor -
Article: Abrogation of anti-HLA antibodies via proteasome inhibition.
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ABSTRACT: Current treatments for autoantibody-mediated diseases (i.e., systemic lupus erythematosus) and alloantibodies (in transplant) are minimally effective. Although they deplete naïve B cells, plasmablasts, and transiently reduce antibody concentrations, they are minimally effective against long-lived, antibody-producing plasma cells. In transplantation, plasma cells produce antibodies directed against human leukocyte antigen (HLA) antigens causing poor allograft survival. We report the first clinical experience with a plasma cell depleting therapy, bortezomib, to abrogate anti-HLA antibodies in transplantation (outside of rejection) in an attempt to improve long-term allograft survival. Eleven patients with anti-HLA alloantibodies were treated with bortezomib. All patients underwent plasmapheresis to aid in removal of antibodies and to determine the effect of bortezomib. Serial measurements of anti-HLA antibody levels were conducted weekly by single antigen bead on Luminex platform. Bortezomib treatment elicited substantial reduction in both donor-specific antibody (DSA) and non-DSA levels. Antibodies were directed against DSA in 8 of 11 cases. Mean time to antibody appearance was 2 months posttransplant. Within 22 days (median) from treatment initiation, 9 of 11 patients' antibody levels dropped to less than 1000 mean fluorescence intensity. Of two patients without successful depletion, all had peak mean fluorescence intensity more than 10,000. At a mean follow-up of approximately 4 months posttreatment, all patients have stable graft function. Minimal transient side effects were noticed with bortezomib in the form of gastrointestinal toxicity, thrombocytopenia, and paresthesias. Bortezomib therapy effectively abrogates anti-HLA antibodies. Hence, removal of antibodies, by proteasome inhibition, represents a new treatment strategy for transplantation and may have benefit in autoimmune-related disease.Transplantation 06/2009; 87(10):1555-61. · 4.00 Impact Factor
