Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.24.3 (collagenase)
18,340 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The study population in this report by Lin et al. was ob/ob mice that have an inherited genetic deficiency of the appetite-suppressing hormone leptin. These mice develop hyperinsulinemia, insulin resistance, and fatty livers. Compared with their lean littermates and wild-type C57BL-6 mice, ob/ob mice have hepatomegaly. In this study, the authors compared three different groups of adult mice (aged 8-10 wk), including male ob/ob C57BL-6 mice, their lean littermates, and wild-type C57BL-6 mice of the same age and sex. The primary purpose of this study was to test the efficacy of metformin for treatment of fatty liver disease in obese, ob/ob mice that develop hyperinsulinemia or insulin resistance and fatty livers. Metformin therapy was found to eliminate fatty liver disease in this model. The potential mechanisms of the action of metformin were the inhibition of hepatic tumor necrosis factor (TNF)alpha and several TNF-inducible responses, which are likely to promote hepatic steatosis and necrosis. In these experiments, ob/ob mice were divided into three treatment groups. Group 1 consisted of eight mice that were treated with metformin and permitted to consume a nutritiously replete liquid mouse diet ad libitum. Mice in group 2 (n = 8) did not receive metformin but were pair-fed the same volume of liquid diet that the mice in the metformin-treated group had consumed on the previous day. Obese ob/ob mice in group 3 (n = 4) and lean mice received no metformin, as with the mice in group 2, but were permitted to consume the liquid diet ad libitum. Liquid diet was given to facilitate accurate daily comparison of food intake among the various treatment groups. All mice were weighed at the beginning of the study and weekly thereafter until killed and then sera, fat, and liver tissues were collected. Tissues were either fixed in buffered formalin and processed from the deceased mice for histology or snap frozen in liquid nitrogen and stored until RNA and proteins were isolated. The feeding protocol was repeated with a second group of 18 ob/ob mice. After 4 wk, hepatocytes were obtained by in situ liver perfusion with collagenase and assayed for cellular adenosine triphosphate (ATP) content. In each experiment, hepatocytes isolated from 3 mice from each treatment group were suspended in a medium and pooled for subsequent analysis to evaluate cell viability, determine the number of obtained cells, and to assay cellular ATP content. These experiments were repeated using another 3 mice from each treatment group, so that analysis of hepatocytes took place from six ob/ob mice in each feeding group.Hepatic steatosis was decreased significantly only in the metformin-treated group. The authors found that metformin's beneficial effect on the fatty liver disease of mice was not due to its ability to constrain hyperphagia, nor due to decreased caloric ingestion, because the daily caloric intakes of the metformin-treated mice and the pair-fed control mice were virtually identical. These caloric intakes were consistently approximately 20% less than that of another obese control group that was permitted to consume diet ad libitum. The authors also observed no significant effect of metformin on serum glucose concentration from fed, ob/ob mice. Metformin is known to reduce hyperinsulinemia by about 40% in both of these obese hyperinsulinemic and insulin-resistant rodent strains. In conclusion, Lin et al. documented that metformin improves fatty liver disease and reverses hepatomegaly, steatosis, and aminotransferase abnormalities in mice. In addition, the authors suggest that metformin might inhibit dieting-induced redistribution of lipid from the liver to adipose tissue depots. In summary, this study identifies a potential treatment for fatty liver disease in humans.
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PMID:Current biochemical studies of nonalcoholic fatty liver disease and nonalcoholic steatohepatitis suggest a new therapeutic approach. 1449 93

The present study was undertaken to investigate whether matrix metalloproteinase (MMP) functions to prevent the occurrence of destructive fibrosis in progressive renal disease. As a sustained release carrier of plasmid DNA, biodegradable hydrogels and microspheres were formulated from cationized gelatin prepared through aminization. Plasmid DNA was released from the cationized gelatin hydrogels as a result of hydrogel degradation. A plasmid DNA including a cytomegalovirus promoter and human recombinant MMP-1 gene (pCMV-MMP) was constructed. Gelatin microspheres incorporating pCMV-MMP as well as phosphate-buffered saline (PBS) with or without pCMV-MMP were injected into the renal subcapsule of C57BL/6 mice, which were intraperitoneally injected with streptozotocin (STZ) to induce diabetes 7 days after operation. The mice were killed 4 weeks after STZ injection to sample their blood and kidneys for biochemical and histological examinations. An immunofluorescence study confirmed that MMP protein was expressed around the renal tissue injected with gelatin microspheres incorporating pCMV-MMP. When applied with cationized gelatin microspheres incorporating pCMV-MMP, the mice showed a level of blood urea nitrogen significantly lower than that of other groups. A reduced content of collagen in the kidneys of mice administered gelatin microspheres incorporating pCMV-MMP was histologically observed. Further, the hydroxyproline assay revealed a significantly decreased content of hydroxyproline in kidney. We conclude that sustained release of MMP-1 gene is a promising prophylactic trial for kidney fibrolysis and dysfunction in the STZ-induced diabetic mouse model.
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PMID:Local delivery of matrix metalloproteinase gene prevents the onset of renal sclerosis in streptozotocin-induced diabetic mice. 1467 37

Alternative approaches to overcome the shortage of donors for liver transplantation may be the use of hepatocytes for bioartificial devices or transplantation. Therefore, the setting-up of new in vitro culture techniques allowing the long-term survival and functional maintenance of hepatocytes represents a formidable challenge. Aim of this study was to obtain a liver homologous acellular matrix (HAM) able to support viability and metabolic functions of rat hepatocytes in primary culture. HAMs were prepared by sequential incubation of rat liver slices in deoxycholic acid and DNase solutions. Dispersed rat hepatocytes were obtained by collagenase digestion and mechanical disaggregation. Isolated hepatocytes were seeded on uncoated and collagen- or HAM-coated tissue culture plastic wells. Cultures were examined by scanning electron microscopy (SEM), and the viability of hepatocytes and their ability to produce albumin and urea were assessed. The viability of freshly dispersed hepatocytes was about 98%. Hepatocytes seeded on HAM exhibited a significantly higher viability and a markedly lower apoptotic rate than those grown on plastic or collagen. Accordingly, albumin and urea nitrogen productions were significantly higher in HAM-cultured hepatocytes. SEM showed that hepatocytes seeded on HAM displayed a clustered organization, and were well anchored to the matrix and morphologically stable. Taken together, these findings indicate that HAM strongly improves viability and functional activity of rat hepatocytes cultured in vitro.
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PMID:Acellular liver matrix improves the survival and functions of isolated rat hepatocytes cultured in vitro. 1537 76

Poly-[N-(2-hydroxyethyl)-L-glutamine] (PHEG) and poly(ethylene glycol) (PEG)-grafted PHEG conjugates of N,N-di(2-chloroethyl)-4-phenylenediamine mustard (PDM) were synthetised. A collagenase-sensitive oligopeptide spacer was selected to link the cytotoxic agent PDM onto the polymeric carrier. First, the oligopeptide-drug conjugate, L-pro-L-leu-gly-L-pro-gly-PDM, was prepared. In a second step, the low molecular weight PDM derivative and PEG-NH(2) were coupled to a N,N-disuccinimidylcarbonate activated PHEG. Dynamic laser light scattering measurements indicated the formation of aggregates. The presence of human serum albumin had no significant effect on the diameter of the conjugates. The hydrolytic stability of the conjugates was investigated in buffer solutions. The conjugates showed an improved stability compared to the parent nitrogen mustard. The enzymatic degradation studies of the polymeric conjugates were performed in the presence of collagenase type IV (Clostridiopeptidase A; EC 3.4.24.3), cathepsin B (EC 3.4.22.1), cathepsin D (EC 3.4.23.5) and tritosomes. Only the bacterial collagenase type IV was able to cleave the spacer releasing free PDM and its peptidyl derivative, gly-L-pro-gly-PDM. The in vitro cytotoxicity of the conjugates was evaluated against HT1080 fibrosarcoma cells and MDA adenocarcinoma cells. All conjugates showed low toxicity towards these cell lines.
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PMID:Synthesis and in vitro evaluation of macromolecular antitumour derivatives based on phenylenediamine mustard. 1566 87

To investigate whether hepatic stellate cells (HSCs) alter their expression of MMPs after exposure to nitrogen oxide intermediate (NOI), a human hepatic stellate cell line, LI90 cells, was stimulated with an NO donor, SNAP, or a peroxynitrite donor, SIN-1, and the culture supernatants were analyzed by gelatin zymography or anti-MMPs immunoblot. Although SIN-1 did not enhance the secretions of MMP-1 and 13, SIN-1 induced the NF-kappaB activation, MT1-MMP expression and the secretion of activated MMP-2 in LI90 cells. These results suggest that peroxynitrite may contribute to the remodeling of the extracellular matrix in liver by activating pro-MMP-2.
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PMID:Peroxynitrite-mediated matrix metalloproteinase-2 activation in human hepatic stellate cells. 1591 83

Smoking induces skin ageing, affects wound healing and inflammatory responses in skin and mucous membranes but the mechanisms behind these adverse effects of smoking are not clear. The objective was to elucidate the mechanisms of smoking-related tissue damage, by comparing the levels of matrix metalloproteinases (MMPs) -2, -9, and -8 in the skin, serum and saliva of smokers and non-smokers. The study population consisted of 47 current smokers and 51 non-smokers, all males of Finnish origin. Skin samples from the upper inner arm were frozen in liquid nitrogen. Levels of MMP-2 and MMP-9 protein in the skin were assessed by zymography and MMP-8 isoforms were determined by Western blotting. From the serum samples, MMP-2 and MMP-9 were assessed by zymography and MMP-8 levels by time-resolved immunofluorometric assay (IFMA). From the salivary samples, MMP-8 levels were analysed by IFMA and MMP-9 levels by capture activity assay. In skin tissue, lower levels of both the pro and active forms of MMP-9 and of the active forms of MMP-8 were found in the smokers compared to the non-smokers. In serum, higher levels of proMMP-2 and proMMP-9 were found in the smokers compared to the non-smokers (P=0.001 and P<0.001, respectively), whereas MMP-8 levels did not differ significantly between the groups. Active forms of MMP-9 and MMP-2 could not be found in serum. In saliva, the amount of total MMP-9 was significantly lower in the smokers (156.0 U/ml) compared to the non-smokers (223.9 U/ml, P=0.032), whereas the levels of MMP-8 or active MMP-9 did not differ significantly between the groups. We conclude that smoking alters the levels of matrix metalloproteinases in skin tissue, serum and saliva, which may affect the turnover of extracellular matrix of skin even though the clinical impact of our findings is not clear.
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PMID:Levels of matrix metalloproteinase-2, -9 and -8 in the skin, serum and saliva of smokers and non-smokers. 1621 64

Oxidative stress generated during islet isolation and transplantation causes islet cell damage. These oxidative injuries are mediated by reactive oxygen species (ROS) and reactive nitrogen species (RNS). MCI-186 is an antioxidant used for clinical treatment of cerebral infarction in Japan. We examined a possible protective effect of MCI-186 on islet cells against oxidative stress. Islets isolated from Sprague-Dawley rats by collagenase P digestion were purified by density gradient centrifugation with Ficoll. Islets were treated with hydrogen peroxide (H(2)O(2); 5 to 250 micromol) in the presence or absence of MCI-186. Cell death was measured by an LDH release assay. Maximum islet cell death was observed at 250 micromol of H(2)O(2). MCI-186 inhibited islet cell death in a dose-dependent manner with significant reduction above 30 micromol. From the results observed we suggest that the antioxidant effects of MCI-186 may prove beneficial to improve the preservation of islet cells.
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PMID:Protective effect of a radical scavenger, MCI-186 on islet cell damages induced by oxidative stress. 1629 28

The ability of the fetal pancreatic islet cells to multiply rendered them a potential tissue for transplantation studies to cure diabetes. A bank of fetal islets could be created with proper storage in liquid nitrogen. The aim of this study is to evaluate the effect of thawing rate and post-thaw culture on the structural and functional integrity of isolated cryopreserved islets of rat fetuses. Fetal rat islets were isolated by the collagenase digestion, cultured for three days, and then cryopreserved using dimethylsulphoxide as cryoprotectant and the step-rate cooling to -40 degrees C before immersing them in liquid nitrogen. The islets were thawed by the slow or fast warming rates using hyperosmolar sucrose solution and then cultured for 1 or 2 days. Insulin and C-peptide contents of the slow thawed islets were higher than those of the control. In the fast thawed islets the contents were similar to those of the control. Insulin and C-peptide release in response to glucose for the slow thawed islets were lower than those of the control and in the fast thawed islets they were similar to that of the control. Histological examination showed irregular periphery and fragmented central part of the large slowly thawed islets, which showed also variable immunohistochemical reaction to anti-insulin serum, ranging from strongly positive reaction to markedly weak reaction. Fast thawed islets showed mostly regular periphery and their reaction to the anti-insulin serum was slightly weaker than that of the control islets. It was concluded that fast thawing and post-thaw culture is much better than slow thawing, as indicated by nearly normal insulin and C-peptide content and release and intact structural integrity.
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PMID:Effect of thawing rate and post-thaw culture on the cryopreserved fetal rat islets: functional and morphological correlation. 1638 64

Encapsulated hepatocyte transplantation is a promising approach to cell transplantation without immunosuppression as an alternative to whole organ liver transplantation. However, the shortage of donor cells for hepatocyte transplantation has not been resolved, and at this critical point, it seems necessary to establish a method of hepatocyte cryopreservation to allow clinical application of hepatocyte transplantation and the development of a bioartificial liver system in the near future. In this study we demonstrated that cryopreserved microencapsulated rat and human hepatocytes can retain their hepatic function and that cryopreserved microencapsulated human hepatocytes transplanted into rat spleen remain viable without immunosuppression. Rat and human hepatocytes were isolated by a collagenase digestion method, and they were microencapsulated with poly-L-lysine. The microencapsulated rat hepatocytes were transferred to culture medium (DMEM containing 10% FBS and 10% DMSO) and immediately frozen in liquid nitrogen. A warm water bath (37 degrees C) was used to thaw the microencapsulated hepatocytes. Hepatic function, drug metabolism, and cell morphology were assessed after 90 days of cryopreservation. After 1 week of cryopreservation, microencapsulated hepatocytes were cultured for up to 2 weeks to assess their hepatic function and morphology. The morphology of human hepatocytes was assessed after 30 days of cryopreservation. Cryopreserved human hepatocytes were transplanted into rat spleen to assess their morphology. Cryopreserved microencapsulated hepatocytes retained their viability and were strongly positive for expression of albumin, OAT2, CYP3A2, and CYP3A9. Two weeks after cultivation, the cryopreserved microencapsulated rat hepatocytes had retained their hepatic function (urea synthesis). Cryopreserved microencapsulated human hepatocytes also mainly survived and retained their hepatic function for at least 30 days after cryopreservation. Moreover, entrapped cryopreserved human hepatocytes also survived and expressed albumin in rat spleen after transplantation. We demonstrated a novel method of long-term cryopreservation of rat and human hepatocytes by using an encapsulation technique, with retention of biological activity and excellent survival of the cryopreserved microencapsulated human hepatocytes transplanted into rat spleen. We believe that this novel approach to hepatocytes cryopreservation provides a new direction in encapsulated cell therapy with the goal of clinical application in the near future.
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PMID:A novel method of cryopreservation of rat and human hepatocytes by using encapsulation technique and possible use for cell transplantation. 1640 71

The level of cAMP in the rats uterus myocytes under the effect of active metabolites of nitrogen and oxygen (NO, NO2- and H2O2) was studied in the conditions of progesterone influence on myocytes. Suspension of cells was selected with the use of collagenase and soy-bean inhibitor of tripsine. The amount of cAMP was determined with the use of standard reagents producted by "Amersham", England. It is established that the basal level of cAMP in cells is 10.4 +/- 0.7 pmol cAMP/mg protein. Incubation of myocytes with forskoline, 0.1 mM, 10 min, resulted in its 3.4 times increase. It testifies to adenilatcyclase activity characteristic of the great majority of cell objects in the uterus myocytes. The level of cAMP in the suspension insignificantly decreases at the protracted affecting with myocytes 10 nM progesterone (1 hour). Donator of nitrogen oxide (0.1 mM sodium nitroprusside) in the presence of progesterone substantially promoted the level of cAMP in cells at the protracted action (17 +/- 3 pmol cAMP/mg protein). Nitrite-anion and hydrogen peroxide in concentration 10 nM (low physiological concentration) did not have the above effect. The results obtained prove that exactly the long-term influence of nitrogen oxide, instead of progesterone, can provide the increase of cAMP level in the uterus myometrium.
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PMID:[Active nitrogen and oxygen metabolites change cAMP level in the uterus myocytes treated by progesterone]. 1656 14


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