Fibrillar collagen plays an essential role in ventricular remodeling, which is a major prognostic factor in various heart diseases. Inflammatory cytokines, including tumor necrosis factor alpha (TNFalpha), have been reported to play a role in various heart diseases and OPC-8212, a quinolinone derivative, has been demonstrated to reduce TNFalpha production. No studies have examined the effects of OPC-8212 on collagen metabolism in connection with inflammatory cytokine and growth factors. Using lipopolysaccharides as a tool to enhance TNFalpha, we examined the effects of OPC-8212 on the expression of type III collagen mRNA [alpha1(III)] in cultured neonatal rat cardiac fibroblasts. We also measured the concentration of TNFalpha and transforming growth factor beta (TGFbeta) in the cultured medium. Northern blot analysis revealed that LPS reduced the expression of alpha1(III) mRNA, and OPC-8212 counteracted this reduction (on average 25% above the reduced level by LPS stimulation). LPS enhanced the TNFalpha concentration in the medium, and OPC-8212 inhibited this enhancement. LPS increased the TGF-beta1 concentration in the cultured medium, while OPC-8212 did not affect this increase. In summary, OPC-8212 counteracted the reduction in type III collagen mRNA expression by LPS accompanied by suppression of the increase in TNFalpha.
The gene of catalytic domain of the protein kinase of RSV-src was cloned into the BamHI cloning site of a translation vector pET-8c which containing T7 RNA polymerase promotor, and transformed BL21 (DE3) pLys S (Studier and Moffatt, 1986). The putative molecular weight of the protein was about 33 kd as evaluated on the basis of its nucleotide size showed the identical mobility in SDS-polyacrylamide gel electrophoresis. However, yield of protein production was not high, probably, because of its instability in Escherichia coli.
A-39-year-old man was admitted to our hospital because of a markedly decreased level of serum cholinesterase found incidentally by a blood test. Detailed examination did not reveal severe liver disease, malignant tumor, infection or organophosphate compound poisoning. Investigation of three generations of his family revealed two homozygous and five heterozygous family members with the cholinesterase deficiency gene E1s indicating familial serum cholinesterase deficiency.(Internal Medicine 31 : 397-399, 1992)
A total of 6 α chains [α1 (IV) to α6 (IV)] have been identified in type IV collagen. We examined the localization of these chains in the myocardium of patients with dilated (DCM) and hypertrophic (HCM) cardiomyopathy. The localization of α1 (IV)-α6 (IV) in biopsy specimens of 5 patients with DCM and 4 with HCM was examined using immunohistochemistry with monoclonal antibodies. Both α1 (IV) and α2 (IV) immunostaining formed thin homogeneous outlines around myocytes in control hearts. In the DCM specimens, α1 (IV) and α2 (IV) immunostaining formed thick and irregular patterns around myocytes. Staining for α1 (IV) and α2 (IV) was also obscrved in some enlarged intercellular spaces. In 3 DCM hearts, moderate staining for α1 (IV) and α2 (IV) was observed in small replacement fibrotic lesions. In large replacement fibrotic lesions, no α1 (IV) or α2 (IV) staining was observed. In the HCM specimcns, α1 (IV) and α2 (IV) staining formed thick homogeneous patterns around myocytes. In the enlargcd intercellular spaces, no α1 (IV) or α2 (IV) staining was observcd. No labeling for α3 (IV)-α6 (IV) was observed in any heart examined. In conclusion, the present results demonstrate that type IV collagen consisting of α1 and α2 chains appears in the fibrotic lesions of DCM, indicating its contribution to the development of fibrotic changes in the myocardium of DCM patients. In contrast, type IV collagen was restricted to the myocyte membrane in the HCM hearts. Fibrotic processes in the intercellular spaces may differ between DCM and HCM hearts.
The type XVII collagen α1 chain has been identified as a component of the type I hemidesmosome, and is thus thought to play a role in extracellular matrix (ECM) maintenance and signal transduction between the cell and the ECM. We examined the expression of type XVII collagen α1 chain mRNA in the mouse heart by Northern blot analysis and determined the sequential changes of its expression in different developmental stages of the heart using the reverse transcriptase-polymerase chain reaction (RT-PCR) method. Northern blotting: Total RNA was extracted from 10 adult mouse hearts by the guanidine/cesium method. Hybridization was performed with mouse cDNA for α1 (XVII) collagen. RT-PCR: Total RNA was extracted from 7 embryos, 4 neonates and 8 adult mice. Reverse transcription was performed using oligo-dT primer and MMLV. Amplification was carried out in α1 (XVII) collagen and glyceraldehyde 3-phosphate dehydrogenase (GAPDH). GAPDH served as an internal control. Northern blotting revealed a 5.6kb signal that was identical to that of the α1 (XVII) of skin and transformed keratinocyte reported previously. The sequences of the PCR products were also identical to those reported. The normalized expression ratios of α1 (XVII) were 0.91±0.20 in the embryonic heart, 0.36±0.20 in the neonatal heart and 0.96±0.21 in the adult heart. In conclusion, we identified the expression of type XVII collagen α1 chain mRNA in the mouse heart, suggesting that the type I hemidesmosome is located in the heart. The results of the RT-PCR at different developmental stages of the heart suggest that type XVII collagen contributes not only to cardiogenesis in the embryonic stage but also to maintenance of architecture and function in the adult heart.
