Role of Periductal and Ductular Epithelial Cells of the Adult Rat Pancreas in Pancreatic Hepatocyte Lineage: A Change in the Differentiation Commitment

Development of pancreatic hepatocytes in adult rats maintained on copper deficient diet containing 0.6% trien (CuDT) has been reported recently. To elucidate the histogenesis of hepatocytes a sequential study was undertaken using morphologic, histochemical, immunochemical, in situ hybridization, and Northern blot analysis. Male F-344 rats weighing 80 to 90 g were fed CuDT for 8 weeks and returned to normal rat chow. Beginning from 4 weeks of copper depletion, there was a progressive loss of acinar cells and by 8 weeks more than 90% of the acinar tissue was lost. During this period, there was an increase in the number of adipocytes in the interstitium, and in the number of interstitial and ductular cells. Morphologic observations were confirmed by immunoblot and Northern blot analysis, in which the amount of pancreatic proteins and their mRNAs decreased between 5 and 8 weeks. During this period, a progressive increase in the level of albumin mRNA was observed. In situ hybridization, performed at 7 weeks of copper deficiency, showed localization of albumin mRNA over interstitial and ductular cells. Pancreatic hepatocytes were identified immediately after the rats were returned to a normal diet and gradually increased in number. The hepatocytes occupied almost 60% of the pancreatic volume by 8 weeks. During the early recovery phase, hepatocytes were identified in ductules as well as in the interstitium. Based on these studies, it is concluded that both the ductular cells and interstitial cells, which resemble oval cells of liver, are capable of transforming into pancreatic hepatocytes and these cells may be considered stem-cell equivalent.     

Androgen receptor YAC transgenic mice carrying CAG 45 alleles show trinucleotide repeat instability

X-linked spinal and bulbar muscular atrophy (SBMA) is caused by a CAG repeat expansion in the first exon of the androgen receptor (AR) gene. Disease-associated alleles (37-66 CAGs) change in length when transmitted from parents to offspring, with a significantly greater tendency to shift size when inherited paternally. As transgenic mice carrying human AR cDNAs with 45 and 66 CAG repeats do not display repeat instability, we attempted to model trinucleotide repeat instability by generating transgenic mice with yeast artificial chromosomes (YACs) carrying AR CAG repeat expansions in their genomic context. Studies of independent lines of AR YAC transgenic mice with CAG 45 alleles reveal intergenerational instability at an overall rate of approximately 10%. We also find that the 45 CAG repeat tracts are significantly more unstable with maternal transmission and as the transmitting mother ages. Of all the CAG/CTG repeat transgenic mice produced to date the AR YAC CAG 45 mice are unstable with the smallest trinucleotide repeat mutations, suggesting that the length threshold for repeat instability in the mouse may be lowered by including the appropriate flanking human DNA sequences. By sequence-tagged site content analysis and long range mapping we determined that one unstable transgenic line has integrated an approximately 70 kb segment of the AR locus due to fragmentation of the AR YAC. Identification of the cis - acting elements that permit CAG tract instability and the trans -acting factors that modulate repeat instability in the AR YAC CAG 45 mice may provide insights into the molecular basis of trinucleotide repeat instability in humans.      

Potentiation of human alpha4beta2 neuronal nicotinic receptors by a Flustra foliacea metabolite

The effects of various Flustra foliacea metabolites on different types of human neuronal nicotinic acetylcholine receptors (nAChRs) expressed in Xenopus oocytes were investigated. Whereas most of the compounds tested had a small blocking effect, one of them, deformylflustrabromine, selectively increased the current obtained in alpha4beta2 receptors when co-applied with acetylcholine (ACh). The current increase was reversible and concentration-dependent. This potentiating effect was still present at saturating concentrations of acetylcholine, and no changes in single-channel conductance or reversal potential were observed, thus suggesting a modification in the gating of alpha4beta2 receptors. Dwell time analysis of single channel records indicates that the mechanism of action of deformylflustrabromine could be both an increase of the opening rate constant and a decrease of the closing rate constant on alpha4beta2 receptors. Thus, deformylflustrabromine may constitute an excellent starting point for the future development of related agents able to potentiate human neuronal nicotinic receptor function.     

Human submandibular-sublingual saliva promotes adhesion of Candida albicans to polymethylmethacrylate.

The purpose of this study was to identify components of saliva that interact with Candida albicans in solution and that may modulate adhesion to dental acrylic (polymethylmethacrylate [PMMA]) surfaces. Saliva-derived pellicles extracted from C. albicans blastoconidia and hyphal-form cells mixed with fresh human submandibular-sublingual saliva (HSMSL) contained predominantly high- and low-molecular-weight mucins (MG1 and MG2, respectively). In contrast, few components from fresh human parotid saliva were adsorbed to yeast cells. Coating PMMA beads with HSMSL significantly enhanced (10-fold) adhesion of both growth forms of C. albicans compared with human parotid saliva (2-fold), suggesting a role for mucins in adhesion. HSMSL-enhanced adhesion was completely abolished by preadsorbing HSMSL with either blastoconidia or hyphal-form cells prior to coating PMMA. However, coating PMMA with purified salivary mucins or the addition of mucin to preadsorbed saliva did not enhance or restore adhesion to levels found with fresh HSMSL. Adhesion assays employing guanidine-treated fresh HSMSL showed a complete lack of Candida binding, suggesting that subjecting HSMSL to dissociating conditions may alter a property of salivary mucins crucial for C. albicans adhesion. Protease and glycosidase treatment of yeast cells significantly reduced adhesion to HSMSL-coated PMMA. In addition, preincubation of C. albicans with mannose and galactose inhibited adhesion to HSMSL-coated PMMA. These results suggest that mucins may play a role in C. albicans adhesion to saliva-coated PMMA and that a glycoprotein on the yeast surface may be involved in these events.     

Trypanosoma cruzi exhibits inter- and intra-strain heterogeneity in molecular karyotype and chromosomal gene location

Molecular karyotypes of 6 strains and 6 clones of Trypanosoma cruzi were determined using orthogonal-field-alternation gel electrophoresis. At least 15 different chromosome-sized DNA molecules, ranging in size from less than 200 kilobase pairs to greater than 2000 kilobase pairs, were resolved for each of the isolates examined. Many of the bands were present in different relative intensities suggesting that the number of individual chromosomes per organism may be considerably higher. Significant inter- and intra-strain differences in molecular karyotype and in the chromosomal locations of the genes for the spliced leader, tubulins, 5S ribosomal RNA and a heat shock protein were found. These marked chromosomal differences among T. cruzi strains and clones may be related to the high degree of phenotypic heterogeneity previously found in this parasite.     

Lack of chemopreventive efficacy of DL-selenomethionine in colon carcinogenesis

Epidemiologic observations and laboratory research have suggested that dietary selenium reduces the risk of colon cancer. Selenium-enriched brewer's yeast as a dietary supplement reduces the incidence of and mortality from cancer of the colon in humans. It is not clear whether the observed inhibitory effect is due to selenomethionine, or to other forms of selenium, or to a mixture of the selenium compounds present in selenium-enriched brewer's yeast. Therefore, bioassay described in this study examined the chemopreventive efficacy of 10 and 15 ppm selenomethionine, equivalent to 3.6 and 5.4 ppm as selenium, against azoxymethane (AOM)-induced colon carcinogenesis. At five weeks of age, groups of male F344 rats were fed diets containing 0 (control diet), 10 or 15 ppm selenomethionine. At seven and eight weeks of age, all rats except those in vehicle-treated groups received s.c. injections of AOM at a dose rate of 15 mg/kg body wt. The rats were maintained on their respective diets for 52 weeks and were then sacrificed. Colon tumors were processed and evaluated histopathologically. Colon tumor incidence and multiplicity were analyzed statistically. No obvious toxic effects were observed following dietary administration of 10 or 15 ppm selenomethionine as indicated by body weight gain. Administration of 10 or 15 ppm selenomethionine had no significant effect on colon tumor incidence and multiplicity. This study suggests that i) selenomethionine lacks chemopreventive efficacy against AOM-induced colon carcinogenesis and ii) other forms of selenium or a mixture of selenium compounds present in selenium-enriched brewer's yeast need to be evaluated for their chemopreventive efficacy.     

Alterations of pancreatic hepatocytes in rats exposed to carcinogens.

In this study, acute and chronic responses of pancreatic hepatocytes induced in F-344 rats by copper depletion-repletion protocol to certain hepatocarcinogens were examined. Administration of a single dose of tannic acid (subcutaneous), aflatoxin B1 (gavage), or lasiocarpine (intraperitoneally) caused characteristic nucleolar segregation in parenchymal cells of liver as well as in pancreatic hepatocytes. Chronic dietary administration of 2-acetylaminofluorene (0.025%) for 12 to 32 weeks led to the development of glutathione S-transferase-P-positive pancreatic hepatocytes in the pancreas. In addition, oval cell proliferation was observed in close association with pancreatic hepatocytes, but not in other areas of pancreas containing residual acinar cells. Oval cells in the pancreas and in the liver that developed in rats after chronic 2-acetylaminofluorene treatment and pancreatic duct cells stained positively with rat liver oval cell marker OV-6 antibodies by immunoperoxidase. These findings indicate that pancreatic hepatocytes respond to carcinogens in a fashion similar to parenchymal cells of liver.     

Intestinal antilectin immunoglobulin A antibody response and immunity to Entamoeba dispar infection following cure of amebic liver abscess

We followed 93 subjects with amebic liver abscess (ALA) and 963 close associate controls at 3-month intervals for 36 months to characterize intestinal and humoral antibody responses to the amebic galactose-inhibitable lectin and to determine whether immunity developed to Entamoeba histolytica or Entamoeba dispar infection following cure of ALA. We found that ALA subjects had a higher prevalence and level of intestinal antilectin immunoglobulin A (IgA) and serum anti-LC3 (cysteine-rich recombinant lectin protein) IgA and IgG antibodies, P < 0.01 and P < 0.05, respectively, compared to controls. The intestinal antilectin IgA antibody response was sustained over a longer time period in ALA subjects (71.8% remained positive at 18 months and 52.6% at 36 months, P < 0.001 compared to 17.6% and 10.3% of controls, respectively). ALA subjects were highly immune to E. dispar infection throughout the study (0% infected at 6 and 36 months, compared to 6.5% and 4.9% of control subjects, respectively, P < 0.05). Upon entry into the study, 6.3% of ALA subjects were infected with E. histolytica; the incidence of new E. histolytica infections in controls (as determined by culture) was too low (1.4%) to determine whether ALA subjects exhibited immunity to new infections. We found that stool cultures every 3 months markedly underestimated the occurrence of new E. histolytica infections, as 15.3% of controls seroconverted after 12 months of follow-up. Unfortunately, under the field conditions present in Durban, South Africa, enzyme-linked immunosorbent assay for detection of lectin antigen in stool yielded unreliable results. In summary, subjects cured of ALA exhibited sustained mucosal IgA antibody responses to the amebic galactose-inhibitable lectin and a high level of immunity to E. dispar infection. Determination of immunity to E. histolytica following cure of ALA will require the use of more sensitive and reliable diagnostic methods.