Magnetoliposomes as Contrast Agents for Longitudinal in vivo Assessment of Transplanted Pancreatic Islets in a Diabetic Rat Model.

Magnetoliposomes (MLs) were synthesized and tested for longitudinal monitoring of transplanted pancreatic islets using magnetic resonance imaging (MRI) in rat models. The rat insulinoma cell line INS-1E and isolated pancreatic islets from outbred and inbred rats were used to optimize labeling conditions in vitro. Strong MRI contrast was generated by islets exposed to 50 µg Fe/ml for 24 hours without any increased cell death, loss of function or other signs of toxicity. In vivo experiments showed that pancreatic islets (50-1000 units) labeled with MLs were detectable for up to 6 weeks post-transplantation in the kidney subcapsular space. Islets were also monitored for two weeks following transplantation through the portal vein of the liver. Hereby, islets labeled with MLs and transplanted under the left kidney capsule were able to correct hyperglycemia and had stable MRI signals until nephrectomy. Interestingly, in vivo MRI of streptozotocin induced diabetic rats transplanted with allogeneic islets demonstrated loss of MRI contrast between 7-16 days, indicative of loss of islet structure. MLs used in this study were not only beneficial for monitoring the location of transplanted islets in vivo with high sensitivity but also reported on islet integrity and hereby indirectly on islet function and rejection.

Quantitative assessment of intestinal first-pass metabolism of oral drugs using portal-vein cannulated rats.

PURPOSE: To evaluate the impact of intestinal first-pass metabolism (Fg) by cytochrome P4503A (CYP3A) and uridine 5'-diphosphate-glucuronosyltransferases (UGT) on in vivo oral absorption of their substrate drugs. METHODS: CYP3A and UGT substrates were orally administered to portal-vein cannulated (PV) rats to evaluate their intestinal availability (Fa · Fg). In the case of CYP3A substrates, vehicle or 1-aminobenzotriazole (ABT), a potent inhibitor of CYP enzymes, was pretreated to assess Fg separately from Fa (Enzyme-inhibition method). On the other hand, since potent inhibitors of UGT have not been identified, Fg of UGT substrate was calculated from total amount of metabolites generated in enterocytes (Metabolite-distribution method). RESULTS: After oral administration of CYP3A substrates in ABT-pretreated rats, the portal and systemic plasma concentrations of the metabolite were nearly the same, indicating almost complete inhibition of intestinal CYP3A-mediated metabolism. Using Enzyme-inhibition method, Fg of midazolam (1 mg/kg) was calculated as 0.71. Additionally, total amount of raloxifene-6-glucuronide generated in enterocytes after oral administration of raloxifene was estimated using Metabolite-distribution method and Fg of raloxifene (0.98 µmol/kg) was calculated as 0.21. CONCLUSIONS: PV rats enabled in vivo quantitative assessment of intestinal first-pass metabolism by CYP3A and UGT. This method is useful for clarifying the cause of low bioavailability.

Assessment of intestinal availability of various drugs in the oral absorption process using portal vein-cannulated rats.

To understand the rate-limiting process of oral drug absorption, not only total bioavailability (F) but also intestinal (F(a) · F(g)) and hepatic (F(h)) availability after oral administration should be evaluated. Usually, F(a) · F(g) of drug is calculated from pharmacokinetic parameters after intravenous and oral administration. This approach is influenced markedly by the estimated value of F(h), which varies with the hepatic blood flow used in the calculations. In this study, portal vein-cannulated rats were used to calculate the F(a) · F(g) of drugs from a single oral dosing experiment without data from intravenous injection. Portal vein-cannulated rats were prepared by a new operative method that enables stable portal vein blood flow. This surgery had no effects on hepatic blood flow and metabolic activity. Our method for calculating F(a) · F(g) was validated by determining both portal and systemic plasma concentration profiles of various drugs possessing different pharmacokinetic properties after oral administration to the portal vein-cannulated rats. Simulation of portal and systemic plasma concentrations by physiologically based pharmacokinetic modeling indicated that the balance of the absorption rate constant (k(a)) and elimination rate constant (k(e)) resulted in different patterns in portal and systemic plasma concentration-time profiles. This study is expected to provide a new experimental animal model that enables identification of the factors that limit oral bioavailability and to provide pharmacokinetic information on the oral absorption process of drugs during drug discovery.

Talin1, a valuable marker for diagnosis and prognostic assessment of human hepatocelluar carcinomas.

BACKGROUND/AIMS: Hepatocellular carcinoma (HCC) is one of the most lethal and prevalent cancers in the human population. Despite its significance, there is only limited understanding of pathological mechanisms and therapeutic options. Talin1, a focal adhesion complex protein that is required for cell adhesion and motility, regulates integrin interactions with extracellular matrix (ECM). In the present study, we aimed to study the possible role of Talin1 in diagnosis and prognosis of HCC. METHODS: Expression of Talin1 protein was detected in normal liver tissues (n=10), HCC tissues (n=32) and adjacent non-cancerous tissues (n=32) by immunohistochemistry and real time PCR. RESULTS: While Talin1 was observed in all tissues, the protein and mRNA expression of Talin1 in HCC tissues was significantly lower than that in the adjacent non-cancerous tissues and normal liver tissues(P<0.05). In addition, the expression of Talin1 in HCCs was significantly correlated with pathological differentiation, integrity of the tumor capsule, portal vein tumor thrombus and tumor size (P<0.05). CONCLUSIONS: Talin1 is possibly involved in the process of the carcinogenesis, infiltration and metastasis of HCC and has potential as a marker for diagnosis and prognostic assessment.

Assessment of the beta 2 adrenoceptor and Ca2+ channel-blocking activity of drugs with the rat portal vein.

The rat portal vein has spontaneous mechanical activity. The effects of beta-adrenoceptor agonists and antagonists and verapamil alone and together on this mechanical activity have been determined. Isoprenaline, but not dobutamine, attenuated the contractile activity. Three successive challenges to isoprenaline produced identical attenuation curves. The responses to isoprenaline were antagonized by propranolol, metoprolol, and ICI 118,551, and the pA2 values, derived by Schild analysis, were 9.12, 6.78, and 9.33, respectively. Thus, the rat portal vein contains predominantly beta 2-adrenoceptors. Verapamil attenuated the contractile activity of the rat portal vein, and this attenuation was not altered by the presence of propranolol at 10(-5) M. The potencies of isoprenaline and propranolol were not altered by the presence of a 30% attenuation of the contractile activity with verapamil at 10(-7) M. Thus, the rat portal vein may be used to determine the potencies of drugs as beta 2-adrenoceptor antagonists and as voltage dependent calcium channel blockers. In addition, the potency of drugs as beta 2-adrenoceptor antagonists on the rat portal vein may be determined in the presence of some voltage dependent calcium channel blockade.