A new liver metastatic and peritoneal dissemination model established from the same human pancreatic cancer cell line: analysis using cDNA macroarray

To elucidate the mechanisms of metastasis, we established two sublines HPC-1H5 with a highly liver metastatic cell line and HPC-1P5a with a highly peritoneal disseminating cell line, which were sequentially selected from the parental pancreatic cancer cell line HPC-1. Using these three cell lines, we investigated several biological properties and mRNA levels of differentially-expressed genes involved in cancer metastasis by cDNA macroarray. Microscopic findings for the three cell lines were the same. The tumorigenicity, in vitro growth ability, motile activity, adhesive activity and the production of IL-8 of metastatic sublines were higher than those of parental HPC-1 cells. Particularly, HPC-1H5 cells showed clearly higher levels of IL-8 expression and tumors of HPC-1H5 cells grew faster and bigger than those of HPC-1P5a cells. In cDNA macroarray analysis of HPC-1H5 cells, 22 genes were up-regulated and 44 genes were down-regulated compared with parental HPC-1 cells. In HPC-1P5a cells, 9 genes were up-regulated and 28 genes were down-regulated compared with parental HPC-1 cells. This study provides a demonstration of global gene expression analysis of pancreatic cancer cells with liver metastasis and peritoneal dissemination. Furthermore, our results provide a new insight into the study of liver metastasis and peritoneal dissemination of human pancreatic cancer. This revised version was published online in July 2006 with corrections to the Cover Date.     

A CASE OF ALPORT'S SYNDROME

A 19-year-old female belonging to a family of Alport's syndrome was autopsied and her kidneys were examined in detail light and electron microscopically. The basement membrane was examined chiefly and the laminated thickening and/or splitting, looseness, irregularity and rail-like appearance of lamina densa were found in the glomerular, Bowman's capsular, tubular and interstitial capillary basement membranes. These findings were strongly suggestive of Alport's syndrome, whether or not the particles are seen in the basement membrane. In addition, Japanese reports on Alport's syndrome (total 48 families) were summarized and renal lesions were examined in comparison. It has been said that the prognosis is worse in the male than in the female, but according to our Investigation on case reports in Japan, the prognosis showed no difference between male and female.     

Specific cellular immune responses to pancreatic antigen in chronic pancreatitis and Sjögren's syndrome

The specific cellular immune response to the partially purified pancreatic antigen was studied by the peripheral blood lymphocyte proliferation assay in patients with chronic pancreatitis, Sjögren's syndrome, and primary biliary cirrhosis. A significant positive result (stimulation index >2.0) was observed in 7 of 21 patients with idiopathic chronic pancreatitis (33%;P<0.05), 6 of 7 patients with Sjögren's syndrome-associated chronic pancreatitis (86%;P<0.0005), and 6 of 11 patients with Sjögren's syndrome (55%;P<0.01), compared to normal controls whose stimulation index was 0.94±0.28 (mean ± SD;n=14; range, 0.56–1.60). On the other hand, patients with alcoholic chronic pancreatitis (17%;n=12), stone-related chronic pancreatitis (0%;n=7), primary biliary cirrhosis-associated chronic pancreatitis (33%;n=3), primary biliary cirrhosis (0%;n=4), systemic lupus erythematosus (17%;n=6), and autoimmune thyroiditis (0%;n=6) showed no significant difference from normal controls. Furthermore, in patients with idiopathic chronic pancreatitis who had positive results, a lymphocyte proliferative response to the pancreatic antigen was observed in T cells, especially in the CD4⁺ T cell subpopulation. These results suggest that the pancreatic antigen plays a role in the pathogenesis of a part of idiopathic chronic pancreatitis and Sjögren's syndrome in association with T cell responses and, also, suggest that autoimmunity may be a possible etiological factor in chronic pancreatitis.     

Developmental expression of carbonic anhydrase-related proteins VIII,X, and XI in the human brain

Three cDNA homologues of carbonic anhydrase with unknown biological functions have been reported: carbonic anhydrase-related proteins (CA-RP) VIII, X, and XI. In the present study, we produced monoclonal antibodies to these CA-RPs and studied their regional and cellular distributions in the human adult and fetal brains by immunohistochemical analysis. In the adult brain, CA-RP VIII was expressed in the neural cell body spreading to most parts of the brain. CA-RP X was expressed in the myelin sheath and its expression was shown in the cytoplasm of cultured tumor cells by immunocytochemical analysis. CA-RP XI was expressed in the neural cell body, neurites, and astrocytes in relatively limited regions of the brain. In the fetal brain, CA-RP VIII and XI were expressed in the neuroprogenitor cells in the subventricular zone as early as the 84th day of gestation and subsequently detected in the neural cells migrating to the cortex. CA-RP X first appeared in the neural cells in the cortex at the 141st day. In the choroid plexus, the epithelial cells gave CA-RP VIII and XI expressions in both adult and fetal brains.From the findings in the present study on the distribution and the developmental expression of CA-RP VIII, X, and XI in the human brain we suggest that these CA-RPs play roles in various biological process of the CNS.     

Effect of magnesium deficiency on ultrastructural changes in coronary arteries of swine

The effects of magnesium (Mg) deficiency on the coronary arteries of 27 Yorkshire swine were studied by light and electron microscopy. The experimental animals were divided into 4 groups which received the following supplements: Group I, basal ration with adequate Mg (540 mg/kg diet), Group II, basal ration with insufficient Mg (270 mg/kg diet) Group III, 10% milk powder with adequate Mg (540 mg/kg diet), Group IV, 10% milk powder with insufficient Mg (270 mg/kg diet). Serum analysis indicated that dietary low Mg supplementation decreased cholesterol levels and increased phospholipid concentrations significantly. The highest magnitude and incidence of intimal thickening were observed in the coronary arteries of Group IV (p less than 0.003). No significant intimal thickening was detected in any of the other groups. Ultrastructural studies revealed a greater frequency of degenerated cells in Group III and IV (p less than 0.01). Numerous calcifications were observed in only Group IV. These data suggest that moderate Mg deficiency can promote atherosclerosis in combination with some atherogenic diet, and that the presence of smooth muscle cell degeneration is important in order for a magnesium deficiency to exert an effect on the coronary artery of swine.     

PRIMARY MALIGNANT FIBROUS HISTIOCYTOMA OF THE ENDOMETRIUM

Malignant fibrous histiocytoma (MFH) has become one of the most common malignancies occurring in soft tissue. To our knowledge, the present case is the first of MFH occurring in the endometrium. The uterus removed from a 47-year-old woman demonstrated a large multinodular endometrial lesion with gross invasive foci in the myometrium and the left oviduct. Microscopically, the endometrial tumor and the invasive lesions were composed of dense sheets of markedly pleomorphic cells consisting of fibroblast-like cells, histiocyte-like cells, foamy histiocytes, benign appearing multinucleated giant cells resembling either osteoclasts or Touton giant cells, and bizarre tumor giant cells. Some of the tumor cells showed phagocytic activities. The tumor cells were oriented in a random or haphazard fashion and classical storiform and fascicular patterns were not observed. The tumor was diagnosed as MFH consisting exclusively of so-called pleomorphic pattern. The patient is alive without evidence of disease, months following total hysterectomy and bilateral salpingo-oophorectomy.     

Cholecystokinin and neurotensin mRNAs are differentially expressed in subnuclei of the ventral tegmental area

Immunohistochemical studies of ventral tegmental area (VTA) neurons indicate that individual cells can contain dopamine as well as the neuropeptide neurotransmitters cholecystokinin (CCK) and neurotensin (NT). We have defined the distribution of the cells expressing the mRNAs encoding these two dopamine cotransmitter peptides in each of the subnuclei of the ventral tegmental area, and quantitated the extent of expression of each gene by using in situ hybridization methods. These studies reveal significant differences in the patterns of expression of each of these two genes within various subdivisions of the VTA. The rostral linear nucleus contained numerous CCK positive cells, some of which appeared to express preproCCK-mRNA at a very high level, but this nucleus contained relatively few NT-expressing cells. The parabrachialis pigmentosus contained numerous NT and CCK positive cells. The paranigralis and interfascicularis nuclei displayed positive CCK cells but with expression at only modest levels. NT cells were very few in these nuclei. The caudal linear nuclei contained the highest number of NT-expressing neurons and these cells expressed very high levels of NT mRNA. The selective distribution of these peptide genes within the VTA subnuclei may have specific consequences. Studies of the connectivity of neurons in the VTA show that the different subnuclei of this region project to several functionally and architectonically different regions of the cerebral cortex and subcortically to nuclei related to the limbic system. Results from our study show very prominent expression of CCK mRNAs in those subnuclei that project heavily to the prefrontal, other cortical areas, and the amygdaloid complex. The NT gene is expressed prominently in those subnuclei of VTA that project heavily to the entorhinal cortex and amygdaloid complex. These results provide support for a differential role for the NT-expressing neurons than that of CCK-expressing neurons of VTA in "reward" mechanisms and in drug-seeking and motivational behavior. These observations could be applied to create working hypotheses and experimental paradigms to test the differential functional activity of the subdivisions of VTA and their potential roles in the pathogenesis and treatment of drug-seeking behavior and other neuropsychiatric disorders.     

Cellular distribution of the NMDA receptor subunit NMDAR1 in fetal ventral mesencephalon transplants in the dopamine-depleted striatum of a rat

Immunohistochemistry was performed to demonstrate the cellular distribution of N-methyl-D-aspartate (NMDA) receptor subunit NMDAR1 in the intrastriatal grafts of a rat model of Parkinson's disease. Unilateral 6-hydroxydopamine (6-OHDA) lesions of the mesostriatal pathway were produced in young adult female rats. Neural transplantation was performed with fetal ventral mesencephalon (VM) tissue (at embryonic day 15) 3 weeks after the 6-OHDA lesions. In the fetal VM in which the tyrosine hydroxylase (TH) immunoreactivity was intensely observed, no NMDAR1 subunit immunoreactivity was detected. Immunopositive cells of NMDAR1 were densely distributed in the intact SNc contralateral to the lesions, in which intense immunoreactivity for TH was observed. In contrast, the cells positive for NMDAR1 in the SNr were scattered. The immunoreactivity for NMDAR1 was markedly decreased in the SNc, but not in the SNr on the lesioned side. Double immunostaining revealed that most TH-positive cells in the SNc showed moderate NMDAR1 immunoreactivity. Within the intrastriatal fetal VM grafts containing TH-positive cells, NMDAR1-positive cells tended to locate homogeneously within the grafts. These were composed of various cell sizes and shapes, but they were mainly medium-sized and aspiny cells. Double immunostaining revealed that a part of the TH-positive cells in the grafts was also immunopositive for NMDAR1. Taken together with our previous studies, it is suggested that both dopaminergic neurons and nondopaminergic neurons in the VM transplants appear to be modified functionally by glutamatergic afferents via various glutamate receptors, including NMDAR1.     

Effects of propiverine hydrochloride (propiverine) on isolated rat and dog urinary bladder]

Propiverine is a drug for the treatment of incontinence and pollakiuria. The effects of propiverine on isolated rat and dog urinary bladder were investigated. At doses of 10(-6)-3 x 10(-5) M, propiverine caused both a rightward shift and inhibition of the maximum response in the acetylcholine (ACh) dose-response curve. The pA2 values for rat and dog urinary bladder were 5.97 and 6.62, respectively. At doses of 10(-5)-10(-5) M, propiverine also dose-dependently inhibited KCl (100 mM)-induced contractions. The IC50 values for rat and dog urinary bladder were 3.9 x 10(-6) M and 3.8 x 10(-6) M, respectively. The pA2 value and the IC50 value of terodiline for rat urinary bladder were 6.08 and 6.6 x 10(-6) M, respectively. In contrast, the pA2 value and the IC50 value of oxybutynin for rat urinary bladder were 7.69 and 4.5 x 10(-6) M, respectively, suggesting that oxybutynin exerts an anti-muscarinic effect at doses at which no discernible anti-KCl effect was observed, whereas propiverine and terodiline exerted both effects at the same doses. The inhibitory effect of drugs on the contraction induced by electrical field stimulation was tested. At a dose of 10(-7) g/ml, tetrodotoxin inhibited the contraction of rat and dog urinary bladder by 76.6% and 92.6%, respectively. Propiverine and verapamil dose-dependently inhibited the contractile response induced by electrical field stimulation at doses of 10(-5) M or more and 3 x 10(-6) M or more, respectively. At these concentrations, a marked anti-KCl effect of the drugs on smooth muscle was observed. On the other hand, atropine caused no inhibition of the contractile response in rat urinary bladder at a dose of 3 x 10(-5) M, and it inhibited the contraction in dog urinary bladder by 14.9% at a dose of 10(-5) M. These findings suggest that although propiverine exhibits both anti-muscarinic and anti-KCl effects in isolated rat and dog urinary bladder, the inhibitory effects of propiverine on the atropine-resistant part of contraction may be mainly due to its anti-KCl effect.