Pancreas and gallbladder agenesis in a newborn with semilobar holoprosencephaly, a case report.

BACKGROUND: Pancreatic agenesis is an extremely rare cause of neonatal diabetes mellitus and has enabled the discovery of several key transcription factors essential for normal pancreas and beta cell development. CASE PRESENTATION: We report a case of a Caucasian female with complete pancreatic agenesis occurring together with semilobar holoprosencephaly (HPE), a more common brain developmental disorder. Clinical findings were later confirmed by autopsy, which also identified agenesis of the gallbladder. Although the sequences of a selected set of genes related to pancreas agenesis or HPE were wild-type, the patient's phenotype suggests a genetic defect that emerges early in embryonic development of brain, gallbladder and pancreas. CONCLUSIONS: Developmental defects of the pancreas and brain can occur together. Identifying the genetic defect may identify a novel key regulator in beta cell development.

Evaluation of a microbiological screening and acceptance procedure for cryopreserved skin allografts based on 14 day cultures.

Viable donor skin is still considered the gold standard for the temporary covering of burns. Since 1985, the Brussels military skin bank supplies cryopreserved viable cadaveric skin for therapeutic use. Unfortunately, viable skin can not be sterilised, which increases the risk of disease transmission. On the other hand, every effort should be made to ensure that the largest possible part of the donated skin is processed into high-performance grafts. Cryopreserved skin allografts that fail bacterial or fungal screening are reworked into 'sterile' non-viable glycerolised skin allografts. The transposition of the European Human Cell and Tissue Directives into Belgian Law has prompted us to install a pragmatic microbiological screening and acceptance procedure, which is based on 14 day enrichment broth cultures of finished product samples and treats the complex issues of 'acceptable bioburden' and 'absence of objectionable organisms'. In this paper we evaluate this procedure applied on 148 skin donations. An incubation time of 14 days allowed for the detection of an additional 16.9% (25/148) of contaminated skin compared to our classic 3 day incubation protocol and consequently increased the share of non-viable glycerolised skin with 8.4%. Importantly, 24% of these slow-growing microorganisms were considered to be potentially pathogenic. In addition, we raise the issue of 'representative sampling' of heterogeneously contaminated skin. In summary, we feel that our present microbiological testing and acceptance procedure assures adequate patient safety and skin availability. The question remains, however, whether the supposed increased safety of our skin grafts outweighs the reduced overall clinical performance and the increase in work load and costs.

Microscopic anatomy of the human islet of Langerhans.

Human islets of Langerhans are complex micro-organs responsible for maintaining glucose homeostasis. Islets contain five different endocrine cell types, which react to changes in plasma nutrient levels with the release of a carefully balanced mixture of islet hormones into the portal vein. Each endocrine cell type is characterized by its own typical secretory granule morphology, different peptide hormone content, and specific endocrine, paracrine, and neuronal interactions. During development, a cascade of transcription factors determines the formation of the endocrine pancreas and its constituting islet cell types. Differences in ontogeny between the ventrally derived head section and the dorsally derived head, body, and tail section are responsible for differences in innervation, blood supply, and endocrine composition. Islet cells show a close topographical relationship to the islet vasculature, and are supplied with a five to tenfold higher blood flow than the exocrine compartment. Islet microanatomy is disturbed in patients with type 1 diabetes, with a marked reduction in beta-cell content and the presence of inflammatory infiltrates. Histopathological lesions in type 2 diabetes are less pathognomonic with a more limited reduction in beta-cell content and occasional deposition of amyloid in the islet interstitial space.

The beta cell population in type 1 diabetes.

Type 1 diabetes is often considered as a disease where more than 90% of the beta cells have been destroyed at clinical onset and where beta cell antigen-driven autoimmune reactivities progressively destroy remaining beta cells as well as newly formed or implanted beta cells. This view will be evaluated in light of histological observations in the pancreas of type 1 diabetic patients and of antibody-positive non-diabetic organ donors.

Computer-aided ultrasonography (HistoScanning): a novel technology for locating and characterizing prostate cancer.

OBJECTIVE: To assess the extent to which prostate HistoScanning (PHS), a new ultrasound-based technology that uses computer-aided analysis to quantify tissue disorganization induced by malignant processes, can identify and characterize foci of prostate cancer compared with step-sectioned radical prostatectomy (RP) specimens. PATIENTS AND METHODS: Between September 2004 and February 2006, 29 men had PHS before their scheduled RP. A three-dimensional ultrasound raw-data file was acquired, and PHS analysed regions of interest (ROI) corresponding to tissue volumes of approximately 0.04 mL. In 13 men the histology was examined on sections of the whole-mount prostate onto which a grid of 5 x 5 mm squares was applied. On a test set of 14 of the 29 patients, PHS analysis was used before knowing the histology results (blinded data), to predict the maximum tumour diameter, focality, laterality and extraprostatic extension (EPE). RESULTS: Identification and characterization by PHS of the index tumour in the 14 patients in the test set correlated closely (r = 0.95, P < 0.001) with the reference test. The concordance in the attribution of multifocality (present/absent), unilateral/bilateral disease between PHS and histology was 100%. EPE as determined by PHS was attributed to all three pT3a pathological specimens in the blinded paired data. In the same set of data, EPE was attributed to one prostate cancer that on pathological inspection was deemed to be organ-confined (pT2b). CONCLUSIONS: PHS has the potential to identify and characterize prostate cancer foci noninvasively. The precision appears to be sufficient to suggest that PHS might be useful as a triage test for men deemed to be at risk of prostate cancer and who wish to avoid prostate biopsy.

Growth and functional maturation of beta-cells in implants of endocrine cells purified from prenatal porcine pancreas.

The development of islet cell transplantation as a cure for diabetes is limited by the shortage of human donor organs. Moreover, currently used grafts exhibit a marginal beta-cell mass with an apparently low capacity for beta-cell renewal and growth. Although duct-associated nonendocrine cells have often been suggested as a potential source for beta-cell production, recent work in mice has demonstrated the role of beta-cells in postnatal growth of the pancreatic beta-cell mass. The present study investigated whether the beta-cell mass can grow in implants that are virtually devoid of nonendocrine cells. Endocrine islet cells were purified from prenatal porcine pancreases (gestation >110 days) and implanted under the kidney capsule of nude mice. beta-Cells initially presented with signs of immaturity: small size, low insulin content, undetectable C-peptide release, and an inability to correct hyperglycemia. They exhibited a proliferative activity that was highest during posttransplant week 1 (2.6 and 5% bromodeoxyuridine [BrdU]-positive beta-cells 4 and 72 h posttransplant) and then decreased over 20 weeks to rates measured in the pancreas (0.2% BrdU-positive cells). beta-Cell proliferation in implants first compensated for beta-cell loss during posttransplant week 1 and then increased the beta-cell number fourfold between posttransplant weeks 1 and 20. Rates of alpha-cell proliferation were only shortly and moderately increased, which explained the shift in cellular composition of the implant (beta-cell 40 vs. 90% and alpha-cell 40 vs. 7% at the start and posttransplant week 20, respectively). beta-Cells progressively matured during the 20 weeks after transplantation, with a twofold increase in cell volume, a sixfold increase in cellular insulin content, plasma C-peptide levels of 1-2 ng/ml, and an ability to correct diabetes. They became structurally organized as homogenous clusters with their secretory vesicles polarized toward fenestrated capillaries. We concluded that the immature beta-cell phenotype provides grafts with a marked potential for beta-cell growth and differentiation and hence may have a potential role in curing diabetes. Cells with this phenotype can be isolated from prenatal organs; their presence in postnatal organs needs to be investigated.

Heterogeneity in distribution of amyloid-positive islets in type-2 diabetic patients.

Amyloid-containing (A+) islets are characteristic for type-2 diabetes (T2D), but their abundance seems variable among patients. It is unclear whether the distribution of A+ islets follows a certain pattern or occurs randomly throughout the pancreatic organ. We investigated the topography of A+ islets in eight pancreata of T2D patients and eight sex- and age-matched non-diabetic subjects. Transversal sections of head, body and tail segments were stained with synaptophysin combined with Congo red to map/quantify islet tissue and amyloid. In the eight T2D pancreata, the overall percentage of A+ islets varied from 4% to 85%. Further analysis in body and tail indicated that peripheral regions exhibited higher percentages of A+ islets than central regions (averages of, respectively, 30% and 17%, P<0.05). Non-diabetic control pancreata also exhibited A+ islets, albeit at a 25-fold lower frequency; a tendency towards higher percentage of A+ islets in peripheral versus central regions was also observed. The higher percentage A+ islets in peripheral regions was associated with a higher density and relative islet over exocrine surface area. These observations on heterogeneity in abundance and distribution of A+ islets need consideration when sampling tissue for studies on human islet amyloidosis. The present methodology allows us to further investigate the susceptibility to amyloidosis of islets in peripheral regions of the pancreas.

Gastrointestinal mesenchymal tumors - immunophenotypic classification and survival analysis.

The current definition of gastrointestinal tumors (GIST) as CD117-positive mesenchymal tumors of uncertain malignant potential fails to include a number of cases with similar histology. In an attempt to improve the classification of these neoplasms, we conducted an immunohistochemical analysis of 244 mesenchymal tumors with histological features of GIST. According to their immunophenotype, the tumors were classified as GISTs, which are characterized by CD117 (c-kit) expression; gastrointestinal CD117-negative CD34 positive stromal tumors (GINST); alpha-smooth muscle actin and/or desmin positive gastrointestinal leiomyogenic tumors (GILT); S-100 and glial fibrillary acidic protein positive gastrointestinal glial/schwannian tumors (GIGT); gastrointestinal neuronal/glial tumors (GINT), which are positive for S-100/glial fibrillary acidic protein plus neuronal/glial markers; and gastrointestinal fibrous tumors (GIFT), which are only vimentin positive. The most common type of tumors were GIST, followed in order of frequency by GINST, GILT, GIGT, GIFT, and GINT. GISTs did not show any preferential location, whereas GINSTs occurred almost exclusively in the stomach and duodenum, and GILTs preferentially in the large intestine. Over a median follow-up period of 71 months, malignant behavior, i.e., metastatic spread, was observed in all tumor types except GINTs. Malignancy was associated with distal gut location, high mitotic activity, large tumor size, and nuclear pleomorphism, though none of these criteria alone discriminated between benign and malignant. Kaplan-Meier analysis of disease-specific survival showed significant differences in the long-term outcome of the newly defined subgroups. We conclude that, despite strong morphological similarities, gastrointestinal mesenchymal tumors are heterogeneous in their immunophenotype and biology.

Omentum is better site than kidney capsule for growth, differentiation, and vascularization of immature porcine ß-cell implants in immunodeficient rats.

BACKGROUND: Rapid revascularization of islet cell implants is important for engraftment and subsequent survival and function. Development of an adequate vascular network is expected to allow adaptive growth of the ß-cell mass. The present study compares omentum and kidney capsule as sites for growth and differentiation of immature ß-cell grafts. METHODS: Perinatal porcine islet cell grafts were implanted in omentum or under kidney capsule of nondiabetic nude rats. Implants were compared over 10 weeks for their respective growth, cellular composition, number and size of ß cells, their proliferative activity, and implant blood vessel density. RESULTS: In both sites, the ß-cell volume increased fourfold between weeks 1 and 10 reflecting a rise in ß-cell number. In the omental implants, however, the cellular insulin reserves and the percent of proliferating cells were twofold higher than in kidney implants. In parallel, the blood vessel density in omental implants increased twofold, reaching a density comparable with islets in adult pig pancreas. A positive correlation was found between the percent bromodeoxyuridine-positive ß cells and the vessel density. CONCLUSIONS: Growth of the ß-cell volume proceeds similarly in the omentum and under the kidney capsule. However, the omentum leads to higher insulin reserves and an increased pool of proliferating cells, which might be related to a more extended vascular network. Our observations support the omentum as an alternative site for immature porcine islet cells, with beneficial effects on proliferation and implant revascularization.

Spontaneous bilateral carotid artery dissection following cervical manipulation.

We report an autopsy case of a bilateral carotid artery dissection, following cervical manipulation by a chiropractor. To establish the etiology of a cervical artery dissection is important in view of possible legal implications and to exclude hereditary disorders, since cervical artery dissection has been linked to several arteriopathies. The underlying arteriopathy in the presented case was an idiopathic cystic medial degeneration. This report emphasizes the role of the pathologist in defining the underlying arteriopathy in carotid artery dissection.

In search of an efficient injection technique for future clinical application of spermatogonial stem cell transplantation: infusion of contrast dyes in isolated cadaveric human testes.

OBJECTIVE: To develop an efficient infusion technique for human spermatogonial stem cell transplantation. DESIGN: A mixture with ultrasonic contrast, computerized tomography (CT) contrast, and Chinese ink was injected into isolated human testes through different sites (the rete testis, the head of the epididymis, the deferent duct, and blind testicular infusion). Ultrasound transducer was used to visualize the injection site and to observe the flow of the mixture injected in the testes. Then, micro-CT scan was used to construct three-dimensional images, allowing the calculation of the testicular volume filled by the mixture. Finally the efficiency of the infusion was evaluated on histologic sections. SETTING: Research laboratory. PATIENT(S): Cadaver testes obtained from autopsied bodies at the department of pathology. INTERVENTION(S): Ultrasound-guided infusion of contrast liquid. MAIN OUTCOME MEASURE(S): Contrast liquid-filled testis volume and presence of ink in seminiferous tubules. RESULT(S): Ultrasonography clearly visualized the flow when seminiferous tubules were injected from the rete testis. No flow was observed when infusions were made either blindly, into the deferent duct, or into the head of the epididymis. On micro-CT no significant differences were observed between the different volumes. After rete testis infusion, ink particles were found in the lumen of the rete testis and in tubules, close and distant from the rete testis. CONCLUSION(S): A single ultrasound-guided injection of 800 µL in the rete testis may provide a promising method to transplant human spermatogonial stem cells in a clinical setting.

Beta-cell replication is increased in donor organs from young patients after prolonged life support.

OBJECTIVE: This study assesses beta-cell replication in human donor organs and examines possible influences of the preterminal clinical conditions. RESEARCH DESIGN AND METHODS: beta-Cell replication was quantified in a consecutive series of n = 363 human organ donors using double immunohistochemistry for Ki67 and insulin. Uni- and multivariate analysis was used to correlate replication levels to clinical donor characteristics and histopathologic findings. RESULTS: beta-Cell replication was virtually absent in most donors, with < or =0.1% Ki67-positive beta-cells in 72% of donors. A subpopulation of donors, however, showed markedly elevated levels of replication of up to 7.0% Ki67-positive beta-cells. beta-Cell replication was accompanied by the increased replication of glucagon-, somatostatin-, and CA19.9-positive cells. Prolonged life support, kidney dysfunction, relatively young donor age, inflammatory infiltration, and prolonged brain death before organ retrieval were all found to be significantly associated with an increased level (> or =90th percentile) of beta-cell replication, with the first three risk factors being independent predictors. Increased beta-cell replication was most often noted in relatively young donors (< or =25 years) who received prolonged (> or =3 days) life support (68%); in contrast, it was rare in donors with a short duration of life support regardless of age (1%). Prolonged life support was accompanied by increased levels of CD68(+) and LCA/CD45(+) infiltration in the pancreatic parenchyma. CONCLUSION: These results indicate that preterminal clinical conditions in (young) organ donors can lead to increased inflammatory infiltration of the pancreas and to increased beta-cell replication.

Cyproheptadine prevents pergolide-induced valvulopathy in rats: an echocardiographic and histopathological study.

Serotonergic drugs, such as pergolide, have been associated with the development of cardiac valvular myxoid thickening and regurgitation in humans and more recently in rats. These effects are potentially mediated by the 5-hydroxytryptamine (5-HT)(2B) receptor (5-HT(2B)R). Therefore, we sought to determine whether cyproheptadine, a 5-HT(2B)R antagonist, might prevent toxic valvulopathy in an animal model of pergolide-induced valvular heart disease. For this purpose, 50 male Wistar rats received daily intraperitoneal injections of pergolide (0.5 mg/kg, n = 14), pergolide (0.5 mg/kg) combined with cyproheptadine (10 mg/kg, n = 12), cyproheptadine (10 mg/kg, n = 12), or no injections (control, n = 12) for 20 wk. Echocardiography was performed blindly at baseline and at 10 and 20 wk followed by pathology. At baseline, no differences between groups were found with echocardiography. At 20 wk, aortic regurgitation was present in all pergolide-treated animals, whereas it was less frequently observed in the other groups (P < 0.0001). For the other valves, this difference was less pronounced. On histopathology, not only aortic but also mitral valves were thicker, myxoid, and exhibited more 5-HT(2B)R-positive cells in pergolide-treated animals compared with the other groups. Moreover, regurgitant aortic and mitral valves were thicker than nonregurgitant aortic and mitral valves. In conclusion, we found that cyproheptadine prevented pergolide-induced valvulopathy in rats, which was associated with a reduced number of 5-HT(2B)R-positive valvular cells. This may have important clinical implications for the prevention of serotonergic drug-induced valvular heart disease.

Dose dependency and reversibility of serotonin-induced valvular heart disease in rats.

Serotonergic drugs may lead to valvular heart disease in humans and more recently also in rats. Although clinical data suggest that dose dependency and reversibility after drug cessation might occur, proof of this is lacking. For that purpose, a total of 106 rats were prospectively enrolled: 22 control animals and 7 groups of 12 rats that received daily subcutaneous serotonin injections (5, 10, 20, 30, 40, 50 and 60 mg/kg respectively) for 12 weeks. At 12 weeks, half of the animals of each group were killed for histological analysis, whereas the remaining rats were further followed (without serotonin injections) for an additional 8 weeks. After 12 weeks of serotonin treatment, aortic and mitral regurgitation (AR, MR) were more frequently observed in the high dose groups (>30 mg/kg) compared to controls. Moreover, aortic and mitral valves were also thicker in the high dose groups compared to controls. After 8 weeks free of serotonin injections, AR and MR were no longer significantly higher than controls. Moreover, aortic and mitral valve thickness had normalized, returning to control levels. In conclusion, this study provides evidence for a dose-dependent valvular toxicity of serotonergic drugs, which appears to be reversible after drug withdrawal.

Differences in baseline lymphocyte counts and autoreactivity are associated with differences in outcome of islet cell transplantation in type 1 diabetic patients.

OBJECTIVE: The metabolic outcome of islet cell transplants in type 1 diabetic patients is variable. This retrospective analysis examines whether differences in recipient characteristics at the time of transplantation are correlated with inadequate graft function. RESEARCH DESIGN AND METHODS: Thirty nonuremic C-peptide-negative type 1 diabetic patients had received an intraportal islet cell graft of comparable size under an ATG-tacrolimus-mycophenolate mofetil regimen. Baseline patient characteristics were compared with outcome parameters during the first 6 posttransplant months (i.e., plasma C-peptide, glycemic variability, and gain of insulin independence). Correlations in univariate analysis were further examined in a multivariate model. RESULTS: Patients that did not become insulin independent exhibited significantly higher counts of B-cells as well as a T-cell autoreactivity against insulinoma-associated protein 2 (IA2) and/or GAD. In one of them, a liver biopsy during posttransplant year 2 showed B-cell accumulations near insulin-positive beta-cell aggregates. Higher baseline total lymphocytes and T-cell autoreactivity were also correlated with lower plasma C-peptide levels and higher glycemic variability. CONCLUSIONS: Higher total and B-cell counts and presence of T-cell autoreactivity at baseline are independently associated with lower graft function in type 1 diabetic patients receiving intraportal islet cells under ATG-tacrolimus-mycophenolate mofetil therapy. Prospective studies are needed to assess whether control of these characteristics can help increase the function of islet cell grafts during the first year posttransplantation.

Screening for insulitis in adult autoantibody-positive organ donors.

Antibodies against islet cell antigens are used as predictive markers of type 1 diabetes, but it is unknown whether they reflect an ongoing autoimmune process in islet tissue. We investigated whether organs from adult donors that are positive for autoantibodies (aAbs) against islet cell antigens exhibit insulitis and/or a reduced beta-cell mass. Serum from 1,507 organ donors (age 25-60 years) was analyzed for islet cell antibodies (ICAs), glutamate decarboxylase aAbs (GADAs), insulinoma-associated protein 2 aAbs (IA-2As), and insulin aAbs. Tissue from the 62 aAb+ donors (4.1%) and from matched controls was examined for the presence of insulitis and for the relative area of insulin+ cells. Insulitis was detected in two cases; it was found in 3 and 9% of the islets and consisted of CD3+/CD8+ T-cells and CD68+ macrophages; in one case, it was associated with insulin+ cells that expressed the proliferation marker Ki67. Both subjects belonged to the subgroup of three donors with positivity for ICA, GADA, and IA-2-Ab and for the susceptible HLA-DQ genotype. Comparison of relative beta-cell area in aAb+ and aAb- donors did not show a significant difference. Insulitis was found in two of the three cases that presented at least three aAbs but in none of the other 59 antibody+ subjects or 62 matched controls. It was only detected in <10% of the islets, some of which presented signs of beta-cell proliferation. No decrease in beta-cell mass was detected in cases with insulitis or in the group of antibody+ subjects.

In vivo model of drug-induced valvular heart disease in rats: pergolide-induced valvular heart disease demonstrated with echocardiography and correlation with pathology.

AIMS: Valvular heart disease (VHD), inducing valvular regurgitation, has been described in carcinoid heart disease and recently in Parkinson's patients treated with pergolide. The aim of this study was to develop an in vivo model of drug-induced valvulopathy with pergolide in rats. METHODS AND RESULTS: Thirty male Wistar rats were given daily injections of either pergolide (0.5 mg/kg intraperitoneally) (n = 8), serotonin (20 mg/kg subcutaneously) (n = 8), or the vehicle only (n = 14) for 5 months. At 20 weeks, echocardiography demonstrated the presence of aortic regurgitation (AR) and/or mitral regurgitation (MR) in serotonin (86% AR, P = 0.0001; 57% MR, P = 0.006) and in pergolide animals (67% AR, P = 0.003; 67% MR, P = 0.003) compared with none in placebo. Pulmonary regurgitation (PR) and tricuspid regurgitation (TR) were found in the serotonin (71% PR, P = 0.19; 100% TR, P = 0.06 vs. placebo), pergolide (100% PR, P = 0.014; 83% TR, P = 0.35 vs. placebo), and placebo groups (36% PR; 57% TR). Tricuspid regurgitant area ratio (jet/atrium), however, was more severe in the serotonin [median 26.5 (range 17-42)%; P = 0.02] and pergolide animals [32 (17-39) %; P = 0.03] compared with placebo [12.5 (5-33)%]. We found a good correlation between valvular regurgitation and histologically assessed valvular thickness. Histological examination revealed the presence of diffusely thickened and myxoid aortic, mitral, and tricuspid valves in serotonin and pergolide animals as seen in VHD. CONCLUSION: We demonstrated, for the first time, that long-term pergolide administration led to VHD in rats. This small animal model will permit further in vivo investigation of drug-induced valvulopathies.

Xenotransplantation of purified pre-natal porcine beta cells in mice normalizes diabetes when a short anti-CD4-CD8 antibody treatment is combined with transient insulin injections.

BACKGROUND: Pre-natal porcine endocrine islet cell grafts were recently shown to contain immature beta cells with a marked potential for growth and differentiation following transplantation, and hence for a progressive and long-term correction of diabetes in immune-incompetent mice. The present study investigates whether these grafts are also capable of correcting hyperglycemia in immune-competent mice receiving a short treatment with anti-CD4-CD8 antibodies. METHODS: Pure endocrine islet cell grafts with 0.5 to 1.0 million beta cells were prepared from pre-natal pigs and transplanted under the kidney capsule of alloxan-diabetic CBA/Ca mice. Survival, growth and function of implanted beta cells were followed by measuring plasma porcine C-peptide and glucose, and graft insulin content at start and at post-transplant (PT) week 35. The effect was studied of a 5-day treatment with non-depleting anti-CD4 YTS177 and depleting anti-CD8 YTS169 antibody, either without or with transient insulin injections. RESULTS: Without antibody treatment, all graft recipients remained porcine C-peptide negative and died. Antibody treatment decreased CD4-expression and percentage CD8 cells for 10 and 18 weeks respectively. It resulted in a 30 week-survival of nine out of 14 graft recipients; all nine had progressively become C-peptide positive but only one proceeded to normoglycemia. When antibody treatment was combined with transient insulin injections, 11 out of 14 graft recipients survived long-term, eight became C-peptide positive and six were normoglycemic at PT week 30. In both groups, surviving recipients exhibited a graft insulin content that was 6- to 9-fold higher than at implantation. CONCLUSIONS: Pre-natal porcine beta cells grow and differentiate when transplanted in diabetic immune-competent mice that have been transiently immune suppressed with anti-CD4 and anti-CD8 monoclonal antibodies. They develop metabolic control when recipients are also transiently treated with insulin injections.

Correlation between beta cell mass and glycemic control in type 1 diabetic recipients of islet cell graft.

Islet grafts can induce insulin independence in type 1 diabetic patients, but their function is variable with only 10% insulin independence after 5 years. We investigated whether cultured grafts with defined beta cell number help standardize metabolic outcome. Nonuremic C-peptide-negative patients received an intraportal graft with 0.5-5.0 x 10(6) beta cells per kilogram of body weight (kg BW) under antithymocyte globulin and mycophenolate mofetil plus tacrolimus. Metabolic outcome at posttransplant (PT) month 2 was used to decide on a second graft under maintenance mycophenolate mofetil/tacrolimus. Graft function was defined by C-peptide >0.5 ng/ml and reduced insulin needs, metabolic control by reductions in HbA(1c), glycemia coefficient of variation, and hypoglycemia. At PT month 2, graft function was present in 16 of 17 recipients of >2 x 10(6) beta cells per kg BW versus 0 of 5 with lower number. The nine patients with C-peptide >1 ng/ml and glycemia coefficient of variation of <25% did not receive a second graft; five of them were insulin-independent until PT month 12. The 12 others received a second implant; it achieved insulin-independence at PT month 12 when the first and second graft contained >2 x 10(6) beta cells per kg BW. Of the 20 recipients of at least one graft with >2 x 10(6) beta cells per kg BW, 17 maintained graft function and metabolic control up to PT month 12. At PT month 12, beta cell function in insulin-independent patients ranged around 25% of age-matched control values. Thus, 1-year metabolic control can be reproducibly achieved and standardized by cultured islet cell grafts with defined beta cell number.

Toxicological data and growth characteristics of single post-feeding larvae and puparia of Calliphora vicina (Diptera: Calliphoridae) obtained from a controlled nordiazepam study.

Larvae of the Calliphora vicina (Diptera: Calliphoridae) were reared on artificial food spiked with different concentrations of nordiazepam. The dynamics of the accumulation and conversion of nordiazepam to its metabolite oxazepam in post-feeding larvae and empty puparia were studied. Analysis was performed using a previously developed liquid chromatography-tandem mass spectrometry (LC-MS/MS) method. This method enabled the detection and quantitation of nordiazepam and oxazepam in single larvae and puparia. Both drugs could be detected in post-feeding larvae and empty puparia. In addition, the influence of nordiazepam on the development and growth of post-feeding larvae was studied. However, no major differences were observed for these parameters between the larvae fed on food containing nordiazepam and the control group. To our knowledge, this is the first report describing the presence of nordiazepam and its metabolite, oxazepam, in single Calliphora vicina larvae and puparia.