James M. Gulizia

Tacrolimus and sirolimus cause insulin resistance in normal sprague dawley rats

Background. Tacrolimus-sirolimus immunosuppression has improved islet graft survival but may affect islet function. Methods. We studied the effects of tacrolimus, sirolimus, or both in normal adult male Sprague Dawley rats. Glucose and insulin response to oral glucose load and pancreas pathology were evaluated after two weeks of daily tacrolimus (1–8 mg/kg/day), sirolimus (0.08–8 mg/kg/day), or low-dose sirolimus (0.08 mg/kg/day) plus tacrolimus (1 mg/kg/day) treatment compared to controls. Results. Tacrolimus and sirolimus each caused dose-dependent hyperglycemia with hyperinsulinemia in response to oral glucose compared to controls, suggesting insulin resistance. At the highest doses of sirolimus, fasting insulin concentrations were high and did not increase with oral glucose suggesting loss of first phase insulin release. The combination of low doses of tacrolimus and sirolimus, at concentrations used in clinical transplantation, resulted in hyperglycemia without hyperinsulinemia after oral glucose administration. The combination of tacrolimus and sirolimus decreased islet size, and increased islet apoptosis more than either medication alone, or controls. Conclusions. In summary, short-term therapy with either tacrolimus or sirolimus causes insulin resistance in normal rats. Combination tacrolimus-sirolimus causes greater islet changes suggesting early islet failure.

Recurrent hepatitis C posttransplant: Early preservation injury may predict poor outcome

Organ cold/warm ischemia is thought to be a risk factor for increased severity of recurrence of hepatitis C (HCV) post liver transplantation. We had noted some HCV patients with preservation injury (PI) to have particularly poor outcomes. Our goal was to determine if PI on biopsy in HCV patients is associated with earlier, more rapidly progressive recurrence or graft and patient survival. Sixty‐nine patients from the University of Nebraska transplant database were included: 23 HCV patients with PI (group = 1), 23 non‐HCV patients with PI (group = 2), and 23 HCV patients without PI (group = 3). Patient groups were matched for gender, age, immunosuppression, and time of transplantation for analysis. No difference in time to recurrence was noted between HCV groups (256 vs. 316 days posttransplant). More patients in group 1 had progression to stage 3 or 4 fibrosis, compared to group 3 (43 vs. 9%, P = 0.02). One‐year survival for groups 1, 2, and 3 was 78, 82, and 100% respectively, whereas 3‐yr survival was 59, 82, and 88% (group 1 vs. group 2 or 3 respectively, P = 0.0055). There was no difference in survival between groups 2 and 3. Patients in group 1 that received antiviral treatment had improved survival, compared to those who did not (P = 0.012). Risk factors for poor survival on univariate analysis included severity of PI (Relative Risk = 2.78, P < 0.001) and donor age of >55 (P = 0.014). Multivariate analysis shows HCV is the most important factor. In conclusion, HCV transplant patients with evidence of early PI on biopsy have poorer survival outcomes than non‐HCV transplant patients with PI or HCV transplant patients without PI. Consideration for antiviral therapy early in the posttransplant course may be warranted in this subset of patients. Liver Transpl 12:134–139, 2006.

Induction of Colonic Aberrant Crypts in Mice by Feeding Apparent N-Nitroso Compounds Derived From Hot Dogs

Nitrite-preserved meats (e.g., hot dogs) may help cause colon cancer because they contain N-nitroso compounds. We tested whether purified hot-dog-derived total apparent N-nitroso compounds (ANC) could induce colonic aberrant crypts, which are putative precursors of colon cancer. We purified ANC precursors in hot dogs and nitrosated them to produce ANC. In preliminary tests, CF1 mice received 1 or 3 i.p. injections of 5 mg azoxymethane (AOM)/kg. In Experiments 1 and 2, female A/J mice received ANC in diet. In Experiment 1, ANC dose initially dropped sharply because the ANC precursors had mostly decomposed but, later in Experiment 1 and throughout Experiment 2, ANC remained at 85 nmol/g diet. Mice were killed after 8 (AOM tests) or 17–34 (ANC tests) wk. Median numbers of aberrant crypts in the distal 2 cm of the colon for 1 and 3 AOM injections, CF1 controls, ANC (Experiment 1), ANC (Experiment 2),and untreated A/J mice were 31, 74, 12, 20, 12, and 5–6, with P < 0.01 for both ANC tests. Experiment 2 showed somewhat increased numbers of colonic mucin-depleted foci in the ANC-treated group. We conclude that hot-dog-derived ANC induced significant numbers of aberrant crypts in the mouse colon.

Outcomes of a patient-to-patient outbreak of genotype 3a hepatitis C†

Between March 2000 and July 2001, at least 99 persons acquired a hepatitis C virus genotype 3a (HCV‐3a) infection in an oncology clinic. This nosocomial HCV outbreak provided an opportunity to examine the subsequent clinical course in a well‐defined cohort. This was a retrospective/prospective observational study of the short‐term significant health outcomes of a large, single‐source, patient‐to‐patient HCV‐3a outbreak. Outbreak patients or their legal representatives consenting to study were enrolled between September 2002 and December 2007. We measured history and physical examinations, medical records, HCV serology, HCV RNA and genotype, liver enzymes, histology, response to antiviral therapy, and liver‐related morbidity and mortality. Sixty‐four of the 99 known HCV‐3a outbreak patients participated. During a 6‐year period, six patients developed life‐threatening complications from liver disease, three died, one received a liver transplant, and two were stable after esophageal variceal banding or diuretic therapy of ascites. Thirty‐three patients underwent antiviral therapy, with 28 achieving a sustained viral remission. One patient acquired HCV‐3a infection sexually from an outbreak patient and was successfully treated. Eleven study patients died of malignancy, including two that had achieved a sustained viral remission after antiviral therapy. Conclusion: Our patient cohort had a nosocomial source and an oncologic or hematologic comorbidity. Compared with previous HCV outcome studies, a patient‐to‐patient HCV outbreak in an oncology clinic exhibited significant morbidity and mortality. Attention is needed to the public health risk of nosocomial HCV transmission, emphasizing infection control, early diagnosis, and therapy. (HEPATOLOGY 2009.)

Colon carcinogenesis in wild type and immune compromised mice after treatment with azoxymethane, and azoxymethane with dextran sodium sulfate

The association between inflammation and the risk of colorectal cancer (CRC) is well documented in animal models and in humans, but the mechanistic role of inflammation in CRC is less well understood. To address this question, the induction of colon tumors was evaluated in (i) wild type (WT) and athymic BALB/c mice treated with the colon carcinogen azoxymethane (AOM) as a single agent, and (ii) in an inflammation model of colon cancer employing AOM and dextran sodium sulfate (DSS) in WT, athymic, TCRβ−/−, TCRδ−/− and TCRβ−/− TCRδ−/− C57Bl/6 mice. The athymic BALB/c mice treated with only AOM developed 90% fewer tumors than the WT mice. The difference in response was not due to metabolic activation of AOM or repair of DNA adducts. In the inflammation model using a standard sequential exposure to AOM followed by DSS treatment, the tumor incidence in WT mice was 58% with 7 adenomas and 6 adenocarcinomas. In contrast, the TCRβ−/−, TCRδ−/− and TCRβ−/−TCRδ−/− C57Bl/6 mice showed adenoma incidences of 10, 33, and 11%, respectively, and none of the immune compromised mice developed adenocarcinomas. When the DSS exposure was increased and the AOM lowered, no difference was observed between WT and TCRβ−/− mice due to an increase in the incidence in the TCR null mice without concomitant increase in the WT mice. No tumors were observed in mice treated with AOM or DSS alone.

Abstract 3446: Induction of colonic aberrant crypts in mice by an apparent N-nitroso compound fraction derived from hot dogs

Proceedings: AACR 101st Annual Meeting 2010‐‐ Apr 17‐21, 2010; Washington, DC Introduction: Nitrite-preserved meat products, e.g., hot dogs, may be a cause of colon cancer. This may occur because such products contain a possibly carcinogenic fraction called “total apparent N-nitroso compounds” (ANC). Feeding such products to humans and mice increased the fecal excretion of ANC. We tested here whether hot-dog-derived ANC could induce colonic aberrant crypts (ABC), a precursor lesion for colon cancer, in mice. Methods: We purified the ANC precursor (ANCP) fraction of commercial hot hogs by adsorption-desorption on silica gel and cation exchange resin, and treated the ANCP with nitrite to convert them to ANC. As a positive control group, adult female A/J mice were given 1 or 3 i.p. injections of 5 mg/kg of azoxymethane (AOM) [Experiment (Exp.) 1]. In Exps. 2 and 3, similar mice were continuously fed the purified hot dog ANC (initial dose, 100 nmol/g diet) in a high-fat or “high-fat stress” diet. In Exp. 2 the ANC dose initially dropped sharply, probably because the ANCP was unstable on thawing, but later in Exp 2 and throughout Exp 3 the ANCP level was stable and was maintained at 100 nmol/g diet. The mice were killed after 8 (for AOM tests) or 24-34 (for ANC tests) weeks of treatment. Feces of the ANC-treated mice contained up to 225 nmol ANC/g, indicating that the ANC reached the colon. The distal 2 cm of each colon was stained with methylne blue, the mucosal surfaces were scanned at 400 x magnification, and ABC were identified and counted. Results: Mean numbers of ANC/colon for 1 and 3 AOM injections, AOM controls, ANC (Exp 2), ANC (Exp. 3) and ANC controls were 28, 76, 14, 20, 14 and 5-8. Standard deviations were <50% of mean values. The effect of ANC was significant by the rank order test (P < 0.01) for both Exps. 2 and 3. Potency of ABC induction per mole ANC was 1.7% (Exp. 2) and 0.4% (Exp. 3) of that for AOM in Exp. 1. In Exp. 3 we noted that the ABC occurred as foci with 1-4 ABC/focus and a mean of 6.8 foci/colon. ANC treatment did not affect the number of foci. Conclusions: Hot-dog-derived ANC induced a significant number of ABC in the mouse colon, but the effect was relatively weak. These results provide a direct link between ANC in a nitrite-preserved meat product and effects in the colon. Partly supported by NIH grant RO1 CA-143460-01. Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3446.