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author:("rio, horatio")
1.  Scaffold Architecture Controls Insulinoma Clustering, Viability, and Insulin Production 
Tissue Engineering. Part A  2014;20(13-14):1784-1793.
Recently, in vitro diagnostic tools have shifted focus toward personalized medicine by incorporating patient cells into traditional test beds. These cell-based platforms commonly utilize two-dimensional substrates that lack the ability to support three-dimensional cell structures seen in vivo. As monolayer cell cultures have previously been shown to function differently than cells in vivo, the results of such in vitro tests may not accurately reflect cell response in vivo. It is therefore of interest to determine the relationships between substrate architecture, cell structure, and cell function in 3D cell-based platforms. To investigate the effect of substrate architecture on insulinoma organization and function, insulinomas were seeded onto 2D gelatin substrates and 3D fibrous gelatin scaffolds with three distinct fiber diameters and fiber densities. Cell viability and clustering was assessed at culture days 3, 5, and 7 with baseline insulin secretion and glucose-stimulated insulin production measured at day 7. Small, closely spaced gelatin fibers promoted the formation of large, rounded insulinoma clusters, whereas monolayer organization and large fibers prevented cell clustering and reduced glucose-stimulated insulin production. Taken together, these data show that scaffold properties can be used to control the organization and function of insulin-producing cells and may be useful as a 3D test bed for diabetes drug development.
PMCID: PMC4086656  PMID: 24410263
2.  Adenocarcinoma arising at ileostomy sites: Two cases and a review of the literature 
Total colectomy with ileostomy placement is a treatment for patients with inflammatory bowel disease or familial adenomatous polyposis (FAP). A rare and late complication of this treatment is carcinoma arising at the ileostomy site. We describe two such cases: a 78-year-old male 30 years after subtotal colectomy and ileostomy for FAP, and an 85-year-old male 50 years after colectomy and ileostomy for ulcerative colitis. The long latency period between creation of the ileostomies and development of carcinoma suggests a chronic metaplasia due to an irritating/inflammatory causative factor. Surgical excision of the mass and relocation of the stoma is the mainstay of therapy, with possible benefits from adjuvant chemotherapy. Newly developed lesions at stoma sites should be biopsied to rule out the possibility of this rare ileostomy complication.
PMCID: PMC4478561  PMID: 26131331
Ileostomy; Carcinoma; Adenocarcinoma; Familial adenomatous polyposis; Inflammatory bowel disease; Complication of ileostomy
3.  Use of Sugar on the Healing of Diabetic Ulcers: A Review 
With the advent of several innovative wound care management tools, the choice of products and treatment modalities available to clinicians continues to expand. High costs associated with wound care, especially diabetic foot wounds, make it important for clinician scientists to research alternative therapies and optimally incorporate them into wound care protocols appropriately. This article reviews using sugar as a treatment option in diabetic foot care and provides a guide to its appropriate use in healing foot ulcers. In addition to a clinical case study, the physiological significance and advantages of sugar are discussed.
PMCID: PMC2956799  PMID: 20920433
diabetic foot ulcers; sugar; wound healing
4.  The Micrograft Concept for Wound Healing: Strategies and Applications 
The standard of care for wound coverage is to use an autologous skin graft. However, large or chronic wounds become an exceptionally challenging problem especially when donor sites are limited. It is important that the clinician be aware of various treatment modalities for wound care and incorporate those methods appropriately in the proper clinical context. This report reviews an alternative to traditional meshed skin grafting for wound coverage: micrografting. The physiological concept of micrografting, along with historical context, and the evolution of the technique are discussed, as well as studies needed for micrograft characterization and future applications of the technique.
PMCID: PMC2909510  PMID: 20663442
diabetic foot ulcers; micrografting; wound healing
8.  Islet xenografts in fully xenogeneic (rat → mouse) chimeras: Evidence for normal regulation of function in a xenogeneic mouse environment 
Surgery  1992;112(2):327-332.
Transplantation of untreated rat bone marrow into mouse recipients conditioned by total-body irradiation results in fully xenogeneic chimerism (rat → mouse). The chimerism is stable for up to 70 months, survival is excellent, and there is no evidence for graft-versus-host disease. We recently reported the long-term survival (> 180 days) of donor-specific pancreatic islet xenografts in these fully xenogeneic chimeras.
Chimeras were prepared and typed for chimerism at 6 weeks, and diabetes was induced by streptozocin injection. Donor-specific pancreatic islets were placed under the renal capsule and recipient blood glucose levels were followed biweekly. The aim of this study was to examine whether the transplanted pancreatic islets exhibited normal function in a xenogeneic environment and assess whether the islet xenografts were not only sufficient to support euglycemia but also regulated in function in response to a glucose challenge.
We report for the first time that donor-specific rat islet xenografts were capable of producing normal basal and peak levels of insulin and responding to a glucose challenge in a manner similar to that of normal mouse islets.
These data indicate that donor-specific rat islet xenografts are functional and regulated normally in fully xenogeneic (rat → mouse) chimeras.
PMCID: PMC3022495  PMID: 1641771
9.  Tacrolimus: A Potential New Treatment for Autoimmune Chronic Active Hepatitis: Results of an Open-Label Preliminary Trial 
Autoimmune chronic active hepatitis (CAH-A) is a chronic liver disease of unknown etiology that is believed to have an autoimmune pathogenesis. The disease is slowly progressive until hepatic failure and portal hypertension develop and either death or liver transplantation occur. Currently, the only widely recognized therapy is the administration of glucocorticoids, which have both anti-inflammatory and immunosuppressive actions. Many patients cannot tolerate such therapy because of the psychiatric, osteoporotic, and weight-enhancing actions of steroids. Tacrolimus (FK 506) is a new macrolide antibiotic that has an immunosuppressive activity that is estimated to be 10–200 times greater than that of cyclosporine. Because of its greater immunosuppressive activity, we have used it in the treatment of 21 patients with autoimmune chronic active hepatitis. Before each subject was treated, a liver biopsy and a panel of hematological, serological, and biochemical parameters were assessed. The Tacrolimus was administered orally at 12-h intervals, and the dose was controlled by monitoring plasma FK trough levels. After 3 months of therapy at an oral dose of 3 mg twice a day, having achieved a median blood level of 0.5 ng/ml, the serum ALT level was reduced by 80%, and the AST level was reduced by 70%. Modest change in the white blood cell count and platelet count were noted. The median BUN level increased from a level of 12 to 18 mg/dl, and the serum creatinine increased from 0.9 to 1.3 mg/dl. These preliminary data demonstrate that: 1) Tacrolimus can be used to successfully treat CAH-A; 2) the response of CAH-A to Tacrolimus treatment is rapid and sustained; and 3) a minor increase in the serum BUN and creatinine levels occurs as a consequence of Tacrolimus treatment. It is anticipated that with continued treatment for periods of 1–2 yr, the natural history of CAH-A will be changed such that hepatic failure and the requirement for liver transplantation may be averted.
PMCID: PMC3022515  PMID: 7537444
10.  Tacrolimus (FK 506), a Treatment for Primary Sclerosing Cholangitis: Results of an Open-Label Preliminary Trial 
Primary sclerosing cholangitis (PSC) is a chronic inflammatory disease of the liver that is characterized by progressive cholestasis and the development of secondary biliary cirrhosis. There is no widely recognized therapy for this disease, although anti-inflammatory agents (steroids), immunosuppressive agents (methotrexate), anti-fibrotics (colchicine), and choleretic agents (ursodeoxycholic acid) have been used in various small series. In the present study, Tacrolimus (FK 506), a new and powerful immunosuppressive macrolide antibiotic, has been used to treat 10 patients with PSC. Each subject had a liver biopsy, ERCP with visualization of the intra-and extrahepatic biliary tree, and a panel of hematological, serological, and biochemical laboratory tests before the initiation of the FK 506 therapy. The FK 506 was administered orally at 12-h intervals and was monitored by serial plasma FK 506 trough levels. After 360 days of treatment, the median serum bilirubin level was reduced by 75%, and the serum alkaline phosphatase was reduced by 70%. Moreover, the serum ALT and AST levels were reduced by 80 and 86%, respectively. No change in the serum level of BUN and creatinine levels occurred as a consequence of the FK 506 treatment These data demonstrate that: 1) FK 506 can be used to treat PSC; 2) the response to FK 506 by patients with PSC is rapid; and, 3) no adverse effect on the serum BUN and creatinine levels was observed. It is anticipated that FK 506 will become an important agent for the treatment of patients with PSC because of its powerful immunosuppressive activity.
PMCID: PMC2982698  PMID: 7532912
Transplantation  1992;53(2):407-414.
This report provides our initial experience in islet isolation and intrahepatic allotransplantation in 21 patients. In group 1, 10 patients underwent combined liver-islet allotransplantation following upper-abdominal exenteration for cancer. In group 2, 4 patients received a combined liver-islet allograft for cirrhosis and diabetes. One patient had plasma C-peptide >3 pM and was therefore excluded from analysis. In group 3, 7 patients received 8 combined cadaveric kidney-islet grafts (one retransplant) for end-stage renal disease secondary to type 1 diabetes mellitus. The islets were separated by a modification of the automated method for human islet isolation and the preparations were infused into the portal vein. Immunosuppression was with FK506 (group 1) plus steroids (groups 2 and 3).
Six patients in group 1 did not require insulin treatment for 5 to >16 months. In groups 2 and 3 none of the patients became insulin-independent, although decreased insulin requirement and stabilization of diabetes were observed.
Our results indicate that rejection is still a major factor limiting the clinical application of islet transplantation in patients with type 1 diabetes mellitus, although other factors such as steroid treatment may contribute to deteriorate islet engraftment and/or function.
PMCID: PMC2967200  PMID: 1738936
Transplantation  1992;53(2):277-283.
We recently reported that reconstitution of lethally irradiated B1O mouse recipients with 40×106 untreated WF rat bone marrow cells resulted in stable fully xenogeneic chimerism (WF rat → B10 mouse). In these animals, the tolerance induced for skin xenografts was highly MHC specific in that donor-specific WF rat skin grafts were significantly prolonged while MHC-disparate third-party xenografts were rapidly rejected (median survival time [MST] = 9 days). We have now examined whether islet cell xenografts placed under the renal capsule of chimeras rendered diabetic with streptozotocin would be accepted and remain functional to maintain euglycemia. Animals were prepared, typed for chimerism at 6 weeks, and diabetes induced with streptozotocin. Donor-specific WF (Rt1Au) islet cell xenografts were significantly prolonged (MST >180 days) in WF → B10 chimeras, while MHC-disparate third-party F344 rat (Rt1Al) grafts were rejected with a time course similar to unmanipulated B10 mice (MST=8 days). The transplanted donor-specific islet cells were functional to maintain euglycemia, since removal of the grafts at from 100 to 180 days in selected individual chimeras uniformly resulted in return of the diabetic state. These data suggest that donor-specific islet cell xenografts are accepted and remain functional in mice rendered tolerant to rat xenoantigens following bone marrow transplantation.
PMCID: PMC2963935  PMID: 1738920
Transplantation  1995;60(12):1421-1425.
On the basis of observations in patients with long-term (28–30 years) renal allograft survival, all of whom had evidence of systemic microchimerism, we began a program of combined simultaneous kidney/bone marrow transplantation. Between 12/14/92, and 10/31/94, 36 kidney transplant recipients received 3–5 × 108 unmodified bone marrow cells/kg; 6 patients also received pancreatic islets, and 7 patients also received a pancreas. The mean recipient age was 39.0 ± 10.8 years, and the mean donor age was 31.8 ± 16.1 years; the mean cold ischemia time was 23.0 ± 9.1 hr. Twenty control patients received kidneys alone, mainly because of refusal by the donor family to consent to vertebral body recovery; 3 of these patients also received a pancreas. The mean recipient age was 47.9 ± 11.7 years, and the mean donor age was 41.5 ± 17.9 years; the mean cold ischemia time was 28.6 ± 6.2 hr. All patients received tacrolimus-based therapy, without radiation, cytoreduction, or induction antilymphocyte preparations. Blood was drawn prior to and at regular intervals after transplantation for detection of chimerism and for immunologic studies. With a mean follow-up of 11.1 ± 5.8 months, all 36 study patients are alive, and 33 (92%) have functioning allografts with a mean serum creatinine of 1.9 ± 1.2 mg/dl and a BUN of 26 ± 9 mg/dl. Graft vs. host disease was not seen in any patient. The incidence of rejection was 72%; 11% of the patients required OKT3 or ATG for steroid-resistant rejection. The incidence of CMV was 14%, and that of delayed graft function was 17%. A total of 18 (90%) control patients are alive, and 17 (85%) have functioning allografts, with a mean serum creatinine of 2.1 ± 1.3 mg/dl, and a BUN of 30 ± 13 mg/dl. The incidence of rejection was 60%, and 10% required OKT3 or ATG. CMV was seen in 15%, and delayed graft function in 20% (P=NS). In the study patients, chimerism was detected in the peripheral blood of 30 of 31 (97%) evaluable patients by either PCR or flow cytometry. In the control patients, chimerism was seen in 9 of 14 (64%) evaluable patients (P<.02). Decreasing donor-specific responsiveness was seen in 6/29 (21%) evaluable study, and 4/14 (29%) evaluable control patients (P=NS). We conclude that combined kidney/bone marrow transplantation is associated with acceptable patient and graft survival, augmentation of chimerism, and no change in the early events after transplantation.
PMCID: PMC2962614  PMID: 8545868
15.  A high throughput live transparent animal bioassay to identify non-toxic small molecules or genes that regulate vertebrate fat metabolism for obesity drug development 
The alarming rise in the obesity epidemic and growing concern for the pathologic consequences of the metabolic syndrome warrant great need for development of obesity-related pharmacotherapeutics. The search for such therapeutics is severely limited by the slow throughput of animal models of obesity. Amenable to placement into a 96 well plate, zebrafish larvae have emerged as one of the highest throughput vertebrate model organisms for performing small molecule screens. A method for visually identifying non-toxic molecular effectors of fat metabolism using a live transparent vertebrate was developed. Given that increased levels of nicotinamide adenine dinucleotide (NAD) via deletion of CD38 have been shown to prevent high fat diet induced obesity in mice in a SIRT-1 dependent fashion we explored the possibility of directly applying NAD to zebrafish.
Zebrafish larvae were incubated with daily refreshing of nile red containing media starting from a developmental stage of equivalent fat content among siblings (3 days post-fertilization, dpf) and continuing with daily refreshing until 7 dpf.
PPAR activators, beta-adrenergic agonists, SIRT-1 activators, and nicotinic acid treatment all caused predicted changes in fat, cholesterol, and gene expression consistent with a high degree of evolutionary conservation of fat metabolism signal transduction extending from man to zebrafish larvae. All changes in fat content were visually quantifiable in a relative fashion using live zebrafish larvae nile red fluorescence microscopy. Resveratrol treatment caused the greatest and most consistent loss of fat content. The resveratrol tetramer Vaticanol B caused loss of fat equivalent in potency to resveratrol alone. Significantly, the direct administration of NAD decreased fat content in zebrafish. Results from knockdown of a zebrafish G-PCR ortholog previously determined to decrease fat content in C. elegans support that future GPR142 antagonists may be effective non-toxic anti-obesity therapeutics.
Owing to the apparently high level of evolutionary conservation of signal transduction pathways regulating lipid metabolism, the zebrafish can be useful for identifying non-toxic small molecules or pharmacological target gene products for developing molecular therapeutics for treating clinical obesity. Our results support the promising potential in applying NAD or resveratrol where the underlying target protein likely involves Sirtuin family member proteins. Furthermore data supports future studies focused on determining whether there is a high concentration window for resveratrol that is effective and non-toxic in high fat obesity murine models.
PMCID: PMC2531115  PMID: 18752667

Results 1-15 (15)