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James R Wright

Publications and source records attributed to James R Wright.

At least 37 records · Page 2Linked to original sources
PubMed2004

Production of transgenic tilapia with Brockmann bodies secreting [desThrB30] human insulin.

BACKGROUND: Tilapia are commercially important tropical fish which, like many teleosts, have anatomically discrete islet organs called Brockmann bodies. When transplanted into diabetic nude mice, tilapia islets provide long-term normoglycemia and mammalian-like glucose tolerance profiles. METHODS: Using site-directed mutagenesis and linker ligation we have "humanized" the tilapia insulin gene so that it codes for [desThrB30] human insulin while maintaining the tilapia regulatory sequences. Following microinjection into fertilized eggs, we screened DNA isolated from whole fry shortly after hatching by PCR. Positive fish were grown to sexual maturity and mated to wild-types and positive Fl's were further characterized. RESULTS: Human insulin was detected in both serum and in the clusters of beta cells scattered throughout the Brockmann bodies. Surrounding non-beta cells as well as other tissues were negative indicating beta cell specific expression. Purification and sequencing of both A-and B-chains verified that the insulin was properly processed and humanized. CONCLUSIONS: After extensive characterization, transgenic tilapia could become a suitable, inexpensive source of islet tissue that can be easily mass-produced for clinical islet xenotransplantation. Because tilapia islets are exceedingly resistant to hypoxia by mammalian standards, transgenic tilapia islets should be ideal for xenotransplantation using immunoisolation techniques.

Amino Acid Sequence↗
PubMed2004

Piscine islet xenotransplantation.

Tilapia, a teleost fish species with large anatomically discrete islet organs (Brockmann bodies; BBs) that can be easily harvested without expensive and fickle islet isolation procedures, make an excellent donor species for experimental islet xenotransplantation research. When transplanted into streptozotocin-diabetic nude or severe combined immunodeficient mice, BBs provide long-term normoglycemia and mammalian-like glucose tolerance profiles. However, when transplanted into euthymic recipients, the mechanism of islet xenograft rejection appears very similar to that of islets from "large animal" donor species such as the very popular fetal/neonatal porcine islet cell clusters (ICCs). Tilapia islets are more versatile than ICCs and can be transplanted (1) into the renal subcapsular space, the cryptorchid or noncryptorchid testis, or intraportally as neovascularized cell transplants; (2) as directly vascularized organ transplants; or (3) intraperitoneally after microencapsulation. Unlike the popular porcine ICCs, BBs function immediately after transplantation; thus, their rejection can be assessed on the basis of loss of function as well as other parameters. We have also shown that transplantation of tilapia BBs into nude mice can be used to study the possible implications of cross-species physiological incompatibilities in xenotransplantation. Unfortunately, tilapia BBs might be unsuitable for clinical islet xenotransplantation because tilapia insulin differs from human insulin by 17 amino acids and, thus, would be immunogenic and less biologically active in humans. Therefore, we have produced transgenic tilapia that express a "humanized" tilapia insulin gene. Future improvements on these transgenic fish may allow tilapia to play an important role in clinical islet xenotransplantation.

Animal Use Alternatives↗
PubMed2004

Evidence that tilapia islets do not express alpha-(1,3)gal: implications for islet xenotransplantation.

BACKGROUND: Cell therapy for diabetes using teleost fish islet tissue has emerged as an intriguing alternative to the use of islet tissue from mammalian pancreases. The islet tissue, called Brockman bodies (BBs), is anatomically distinct from the pancreatic exocrine tissue and can be easily identified and isolated. Islets harvested from Nile tilapia (Oreochromis niloticus), when transplanted into streptozotocin-diabetic nude mice, produce long-term normoglycemia and achieve mammalian-like glucose tolerance profiles. We asked whether tilapia express the alpha-(1,3)gal epitope, the immunodominant target of human xenogeneic responses. METHODS AND RESULTS: Immunostaining with the alpha-(1,3)gal-specific IB4 lectin on tilapia BB, liver, heart, spleen, and head kidney was negative, as was staining with murine anti-alpha-gal-specific monoclonal antibodies. Absence of alpha-gal-specific binding of IB4 or murine anti-gal mAbs to dispersed BBs was confirmed by fluorescent-activated cell sorter analysis. Tilapia BB cell membranes failed to reduce binding of anti-alpha-(1,3)gal-specific mAb in an enzyme-linked immunosorbent assay (ELISA) inhibition assay, while porcine and murine tissue lysates did. Tilapia BB cell lysates were shown to be devoid of alpha-1,3 galactosyltransferase activity by ELISA. Transplantation of tilapia BBs into diabetic alpha-gal knockout (gal KO) mice was not associated with accelerated xenograft rejection when compared with wild type control recipients (mean survival time 6.5 days vs. 7.2 days). Tilapia BBs failed to induce a rise in anti-gal IgG and IgM titers in gal KO mice, while the transplant of wild type mouse islets into gal KO mice caused a significant rise in anti-gal IgG and IgM antibodies. CONCLUSIONS: We conclude that tilapia BBs are devoid of alpha-gal expression, and may offer an alternative to swine as a donor species for islet xenotransplantation.

Animals↗
PubMed2003

Islet transplantation in the discordant tilapia-to-mouse model: a novel application of alginate microencapsulation in the study of xenograft rejection.

BACKGROUND: Tilapia islet xenograft rejection is characterized by infiltration with macrophages (Mphis), eosinophils (Ephis), and T lymphocytes. The presence of these cells indicates they contribute to rejection; therefore, an attempt was made to assess their role through host immunomodulation. METHODS: Tilapia islet cells were transplanted under the kidney capsule of streptozotocin diabetic Balb/c mice, which were then treated with one of several immunomodulatory regimes targeting Mphis, Ephis, or T cells. Mphis were depleted using either silica or liposome-entrapped Cl2MDP. Ephi migration was blocked using monoclonal antibodies (mAbs) targeting interleukin (IL)-4 or IL-5. T-cell function was altered with mAbs targeting CD3, CD4, or CD8. Finally, T helper (Th)1 and Th2 activity was altered by depleting essential Th1 or Th2 cytokines with mAbs or by promoting a Th1 response with the injection of exogenous IL-12. The effects of antibody-mediated immunomodulation on graft survival were initially screened by cotransplanting alginate-encapsulated, mAb-secreting hybridoma cells into the peritoneal cavity at the time of islet transplantation. Significant prolongation was then confirmed using purified antibodies injected at the time of islet transplantation. Rejected grafts were examined histologically, and immunohistochemistry was used to assess the cellular infiltrates for each of the treatment groups. RESULTS: Modulation of Mphis and Ephis alone did not significantly delay functional rejection of tilapia islet grafts (maximal mean graft survival time [mGST]=7.1+/-1.7 and 9.4+/-3.4, respectively) compared with untreated controls (mGST=8.2+/-1.0). Treatment of transplanted animals with antibodies against CD3 or CD4 significantly promoted graft survival (maximal mGST=16.3+/-5.8 and 34.0+/-11.6, respectively), whereas targeting CD8 and Th1 and Th2 cytokines showed no prolonging effect (maximal mGST=7.8+/-2.9 and 9.5+/-4.3, respectively). CONCLUSION: Our results indicate that rejection in the tilapia-to-mouse model follows a pattern similar to other models of discordant islet cell xenotransplantation.

Alginates↗
PubMed2003

Xenogeneic milieu markedly remodels endocrine cell populations after transplantation of fish islets into streptozotocin-diabetic nude mice.

It is unknown whether irrelevant foreign endocrine products secreted by xenografts would be biologically active and potentially harmful to recipients; even if entirely inert, continuous production might result in harmful circulating antigen-antibody complexes. We examined the fate of such a product using a fish (tilapia)-to-mouse islet xenograft model. Teleost fish islets, like mammalian islets, are composed primarily of cells producing insulin, glucagon or somatostatin; however, teleost fish have two different populations of somatostatin (SST) producing delta cells, one producing SST-14, a 14 amino acid SST identical to mammalian SST, which is derived from the pre-proSST-I gene which is present in all vertebrates, and the other a "large" (i.e. 22 to 28 amino acid) SST derived from a pre-proSST-II gene, which is not found in mammals. In contrast to 'large' SST, which has no mammalian homolog, teleost fish insulins, glucagons and SST-14 exhibit significant biological activity in mammals. Tilapia islets were transplanted under the kidney capsules of streptozotocin-diabetic nude mice, and mice with functioning grafts were killed at various times after transplantation. Serial sections of graft-bearing kidneys were stained by immunoperoxidase for insulin, SST-14, SST-25 and glucagon positive cells, and the areas of each cell type in the graft were measured using image analysis. Sections of untransplanted tilapia islets (both in situ and after harvest/culture) were also immunostained and measured as controls. Xenotransplantation of fish islets into diabetic nude mice resulted in the rapid degeneration and near total loss of SST-25+ cells, as well as a marked redistribution of the proportions of the remaining endocrine cell types. The proportions of cell types in the grafts gradually changed from a piscine pattern to that of mammalian islets.

Animals↗
PubMed2002

Almost famous: E. Clark Noble, the common thread in the discovery of insulin and vinblastine.

Clark Noble was one of the first members of the University of Toronto insulin team and came within a coin toss of replacing Charles Best as Frederick Banting's assistant during the summer of 1921. Noble performed important early studies helping to characterize insulin's action, and he co-authored many of the original papers describing insulin. Because mass production of insulin from livestock pancreata had proved elusive throughout 1922, J.J.R. Macleod hired Noble during the summer of 1923 to help him test and develop a new method for producing commercial quantities of insulin that Macleod believed would revolutionize insulin production. However, commercial production of insulin from fish proved impractical and was dropped by 1924, as methods to produce large quantities of mammalian insulin had improved very rapidly. Noble later played a small but critical role in the most important Canadian contribution to cancer chemotherapy research: the discovery of vinca alkaloids by his brother Robert Laing Noble. Although one might expect that a physician involved in 2 of Canada's most important medical discoveries during the 20th century must be famous, such was not Clark Noble's fate. He died without so much as an obituary in CMAJ.

Canada↗
PubMed2002

Factors that influence the recruitment of patients to Phase III studies in oncology: the perspective of the clinical research associate.

BACKGROUND: The multiple determinants of a patient's decision to enter into a clinical trial have been explored largely from the perspectives of patients and their physicians. Little research has involved clinical research associates (CRAs) formally, despite their central role in the process of recruitment. The current study was initiated to explore the factors that influence the decision of patients with cancer regarding clinical trial entry, specifically from the perspective of the CRA. METHODS: Two focus groups of CRAs from the Hamilton Regional Cancer Center were organized. A skilled facilitator guided both groups through exploratory and subsequent confirmatory phases of discussions, which were audiotaped for review and coding using a process of consensus employing intercoder triangulation. RESULTS: The two groups identified a number of factors that they believed influenced the recruitment process. Numerous physician and patient factors were reaffirmed, such as the impression of the scientific merit of a study or the sense of personal benefit, respectively. More uniquely, CRAs identified information transfer within the informed consent process as a major aspect of their specialized role. It was believed that full disclosure of information, in terms of both the content and the techniques and styles of delivery, was an important predictor of recruitment success. The groups quickly reached consensus on which factors they believed were the most important overall with respect to influencing study recruitment. CONCLUSIONS: CRAs appear to have a unique role in the process of recruiting patients to active clinical trials. They believe that they have an important influence on recruitment success. Further research to validate this impression is required, because, ultimately, a greater understanding of the relative roles of physician and patient factors and, potentially, CRA factors will be important in the development of ethical and supportive strategies to optimize the recruitment of patients with cancer into randomized clinical trials.

Attitude of Health Personnel↗
PubMed2002

Eosinophilic/T-cell chorionic vasculitis.

Chorionic vasculitis is the hallmark of a fetal response in chorioamnionitis. There are five highly characteristic findings: (1) leukocyte migration is not concentric but rather radiates toward the infected amniotic fluid; (2) the infiltrate is primarily neutrophils; (3) multiple chorionic vessels, first veins and then arteries, are usually involved; (4) the infiltrate never extends into the vasculature of stem villi; and (5) it is rare in the absence of chorioamnionitis (or its precursors). Here we describe a new form of chorionic vasculitis characterized by an infiltrate composed primarily of eosinophils and CD3+ T lymphocytes that very focally involves a single chorionic vessel (artery or vein), that radiates away from the amniotic fluid (i.e., toward the intervillous spaces), and that may extend into the stem villous vasculature; this lesion occurs in the absence of any evidence of chorioamnionitis. During the past 7+ years, using accepted placental review criteria, we have examined 7104 placentas and identified 14 cases of eosinophilic/T-cell chorionic vasculitis (or related lesions). Although the frequency of diagnosis in the placentas examined was 0.197%, its true incidence cannot be estimated because of its very focal nature and the limited nature of placental disk sampling. Its etiology and significance are unknown, but it may represent a focal immune-mediated vasculitis.

Antigens, CD↗
PubMed2002

From ugly fish to conquer death: J J R Macleod's fish insulin research, 1922-24.

Fish insulin research had a very short heyday. Throughout most of 1922, production of insulin from livestock was difficult, erratic, and expensive. Although fish insulin was easy to extract and seemed to be a brilliant and logical solution to the shortage of insulin, collection of fish islets was a logistical nightmare. By the end of 1922, a scientist at the pharmaceutical company Eli Lilly in Indianapolis, IN, USA had developed a way to concentrate and purify insulin by isoelectric precipitation. As a result of this breakthrough, the scales began to tip heavily in favour of livestock insulin. Nevertheless, research on commercial production of fish insulin continued for another 18 months but was finally abandoned.

Animals↗
PubMed2002

Liposomal encapsulation significantly enchances the immunosuppressive effect of tacrolimus in a discordant islet xenotransplant model.

BACKGROUND: Encapsulation of tacrolimus (TAC) in a lipid bilayer to form liposome-encapsulated tacrolimus (LTAC) alters the biodistribution profile, half-life, and efficacy in organ allotransplantation models. LTAC has not been applied to either cell transplantation or xenotransplantation. METHODS: To test the efficacy of LTAC in a discordant islet xenograft model, tilapia (fish) islets were transplanted under the left kidney capsules of streptozotocin-diabetic Balb/c mice. Recipient mice (groups I-VI) were treated with: I, untreated; II, empty liposomes; III, TAC (2 mg/kg/day); IV, TAC (5 mg/kg/day); V, LTAC (2 mg/kg/day); or VI, LTAC (5 mg/kg/day); all treatments were for 35 days or until rejection (i.e., two glucose measurements >200 mg/dl). Graft-bearing kidneys were removed for histology after rejection. RESULTS: Mean graft survival time (mGST) for control groups I and II were 6.7+/-1.4 (n=6) and 7.5+/-1.3 days (n=4), respectively. Daily TAC treatment at 2 mg/kg/d (III) did not prolong graft function (mGST=7.7+/-1.6; n=6) although 5 mg/kg/day (IV) produced minimal prolongation to 12.8+/-4.8 days (n=12). Treatment with LTAC at 2 mg/kg/day (V) significantly prolonged mGST to 26.6+/-4.9 (n=5); however, all recipients rejected during treatment (i.e.,<35 days). LTAC at 5 mg/kg/day (VI) further prolonged mGST to 39.9+/-11.8 days (n=12) with only one mouse rejecting before day 35. Histologically, at the time of functional rejection, grafts were generally either totally or partially effaced by mononuclear cells, eosinophils, and fibrosis. In groups VI, islet grafts removed from two mice that died while they were normoglycemic and from a mouse terminated while it was normoglycemic at day 36 were viable, well-granulated, and free from cellular infiltration. The group VI grafts examined at rejection (i.e., 1-2 weeks after discontinuing LTAC) were generally totally obliterated and were in two instances associated with nodular aggregates of atypical lymphocytes resembling posttransplant lymphoproliferative disorder. CONCLUSIONS: LTAC is the most potent immunosuppressive compound we have tested in our discordant fish-to-mouse islet xenograft model; however, toxicity is an issue at high doses.

Animals↗
PubMed2002

T-cell/periodic acid-Schiff stain: a useful tool in the evaluation of tubulointerstitial infiltrates as a component of renal allograft rejection.

Distinguishing between tubulitis and tubulointerstitial mononuclear cell infiltrates and determining the severity of tubulitis are critical components of diagnosing and grading renal allograft rejection using the 1993 Banff schema, the revised 1997 Banff schema, or the Cooperative Clinical Trials in Transplantation grading system. We describe a novel staining method, the T-PAS stain (CD3 and periodic acid-Schiff), which removes some of the subjectivity in the evaluation of tubulointerstitial infiltrates in renal allograft biopsies. The method simply combines two routine stains, immunoperoxidase staining for T cells (CD3) and periodic acid-Schiff (PAS) staining for tubular basement membrane, on the same section.

Basement Membrane↗
PubMed2002

Cutaneous ciliated cyst of the abdominal wall: a case report with a review of the literature and discussion of pathogenesis.

A cutaneous ciliated cyst is a rare lesion typically found on the lower extremity of young girls shortly after puberty. Here, we report a case involving a previously unreported site (i.e., abdominal wall) in a 14-year-old girl. We also describe immunohistochemical and ultrastructural findings, review and analyze the world literature, and offer insights as to the pathogenesis of this lesion.

Abdominal Muscles↗
PubMed2002

Human beta cells are exceedingly resistant to streptozotocin in vivo.

Streptozotocin (STZ) causes beta cell death in rodents via the mechanism of DNA damage precipitating poly(ADP-ribose) synthetase activation followed by lethal nicotinamide adenine dinucleotide depletion. It is unclear whether humans are susceptible to this mechanism. Islets were isolated from STZ-sensitive (CD1 mice and Lewis rats) and resistant [fish (tilapia)] species and from man and then were transplanted into diabetic nude mice under the kidney capsule. Normoglycemic recipients with normal glucose tolerance tests on d 30 were injected with increasing iv doses of STZ and their plasma glucose levels followed for 5 d; glucose tolerance tests were repeated on nondiabetic mice. Mice were then killed; grafts and native pancreata were examined. Based upon three criteria (i.e. nonfasting plasma glucose levels, glucose tolerance tests, and islet histology), the following observations were made: 1) Recipients of rat islets were resistant to 25 mg/kg but were uniformly diabetic at doses of 50 or 75 mg/kg. 2) Recipients of mouse islets were resistant to 75 mg/kg but were uniformly diabetic at 150 or 200 mg/kg. 3) Recipients of the fish islets were resistant to 300, 400, and 450 mg/kg. 4) Recipients of human islets were resistant to 100, 200, 300, 400, and 450 mg/kg. The results in recipient mice bearing long-term rat, mouse, or fish islet grafts were the same as previously published dose-response data for each donor species. We extrapolate from our results based on human islet grafts in mice that human beta cells are exceedingly resistant to STZ.

Animals↗