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Objective To observe the inhibitory effect of caudal vein injection of senescence marker protein 30（SMP30） on myocardial injury in septic mice and to explore its mechanism. Methods Sixty male C57BL/6 mice were randomly divided into the Con+empty vector group, LPS+empty vector group, and LPS+SMP30 group, with 20 mice in each. Mice in the Con+empty vector group and LPS+empty vector group were injected with adeno-null associated virus vector via the tail vein, and mice in the LPS+SMP30 group were injected with SMP30 adeno-associated virus vector via the tail vein. After 2 weeks of adeno-associated virus vector injection, mice in the LPS+empty vector group and LPS+SMP30 group were intraperitoneally injected with lipopolysaccharide（LPS） to establish the sepsis models, while mice in the Con+empty vector group were given intraperitoneal injection of the same volume of normal saline. Left ventricular ejection fraction（LVEF） and left ventricular fraction shortening（LVFS） were measured by echocardiography at 24 h after LPS injection.Serum and heart tissues were collected after the mice were killed, and serum levels of creatine kinase-MK（CK-MB), lactic dehydrogenase（LDH） and cTnI, and myocardial levels of malondialdehyde（MDA), superoxide dismutase（SOD） and glutathione peroxidase（GSH-Px） were detected by ELISA. TUNEL staining was used to detect cardiomyocyte apoptosis.Macrophage infiltration was detected by immunofluorescence staining. The myocardial reactive oxide species（ROS） production was detected by dihydroethidium fluorescent probe. Interleukin（IL）-1β, tumor necrosis factor（TNF）-α and IL-6mRNA levels were detected by qRT-PCR. The protein expression levels of myocardial tissues SMP30, silent information regulator 1（SIRT1), Ac-NF-κB p65, and Ac-FOXO1 were detected by Western blotting. Results Compared with the Con+empty vector group, the values of LVEF and LVFS decreased, the levels of serum CK-MB, LDH and cTnI increased, the apoptosis rate increased, the myocardial macrophage infiltration was more obvious, the mRNA levels of IL-1β, IL-6, TNF-α increased, the productions of ROS and MDA increased, the activities of SOD and GSH-Px decreased, the expression of SIRT1 down-regulated, and the expression levels of Ac-NF-κB p65 and Ac-FOXO1 increased in the LPS+empty vector group（all P＜0. 01). Compared with LPS+empty vector group, the values of LVEF and LVFS increased, the levels of serum CK-MB, LDH and cTnI decreased, the apoptosis rate decreased, the myocardial macrophage infiltration decreased, the mRNA levels of IL-1β, IL-6, TNF-α decreased, the productions of ROS and MDA decreased, the activities of SOD and GSH-Px increased, the expression of SIRT1 increased, and the expression levels of Ac-NF-κB p65 and AcFOXO1 decreased in the LPS+SMP30 group（all P＜0. 01). Conclusion The caudal vein injection of SMP30 can up-regulate SMP30 expression in mice with sepsis, inhibit myocardial inflammatory response and oxidative stress level, thereby alleviating myocardial injury, and its mechanism may be related to activating the SIRT1 signaling pathway. [ABSTRACT FROM AUTHOR]

Human senescence marker protein 30 (huSMP30) has been characterized as a multifaceted protein consisting of various enzymatic and cellular functions. It catalyzes the interconversion of L-gulonate and L-gulono-γ-lactone in the ascorbate biosynthesis pathway. Therefore, we hypothesized that it could be a potential anti-biofilm agent against pathogenic bacteria due to its lactonase activity. In order to corroborate this, the huSMP30 was recombinantly expressed, purified, and analyzed for its ability to inhibit Mycobacterium smegmatis biofilm formation, which showed a concentration-dependent inhibition as compared to the untreated control group. Further, in silico analysis was performed to redesign the huSMP30 with enhanced lactonase activity. Molecular docking analysis of the huSMP30 and lactone substrates facilitated the selection of three single amino acid substitutions (E18H, N154Q, and D204V), which were created using a PCR-based site-directed mutagenesis reaction. These mutant proteins and the wild-type huSMP30 were purified, and the effects on the enzymatic activity and biofilm formation were studied. The mutants E18H and D204V showed non-significant effects on specific lactonase activity, catalytic efficiency, and anti-biofilm property; however, the mutant N154Q showed significant improvement in the specific lactonase activity, catalytic efficiency, and inhibition in the biofilm formation. The protein stability analysis revealed that the wild-type huSMP30 and its designed mutants were stable at 37 °C for up to 4 days. In conclusion, the anti-biofilm property of the huSMP30 has been established, and an engineered version, N154Q, inhibits biofilm formation with greater efficiency. Human SMP30 is a versatile protein with multiple cellular and enzymatic functions, however, its anti-biofilm potential has not been explored. Our work presents the method to produce soluble and active huSMP30 in the E. coli expression system and establishes its role as an anti-biofilm agent against Mycobacterium smegmatis owing to its lactonase activity. Our results provide support for the future advancement of huSMP30 as a potential anti-biofilm agent targeting pathogenic Mycobacterium species. [ABSTRACT FROM AUTHOR]

Background/Aim: Chronic kidney disease (CKD) is one of the most common causes of mortality in wild nondomestic felidae. The molecular mechanism regulating renal fibrosis in nephropathy is not fully understood especially in the felidae. This study aimed to elucidate senescence marker protein 30 (SMP30) expression patterns and its relationship with epithelial-mesenchymal transition (EMT) by immunostaining in two necropsied Siberian tigers (Panthera tigris altaica) with CKD. Materials and Methods: Two kidney samples from male Siberian tigers were fixed and tissue sections were stained for histopathological assay. Results: In CKD, renal tubular epithelial cells lost their tubular structures surrounded by severe interstitial fibrosis and were detached from the basement membrane. These damaged cells resembled the morphology of mesenchymal cells and showed much lower SMP30 expression compared with intact tubular epithelial cells. These cells also expressed vimentin, which is specifically expressed by mesenchymal cells, and through double staining, it was observed that vimentin was expressed in the tubular epithelial cells where SMP30 was not expressed. In addition, double-positive expression of pan-cytokeratin (pan-CK) and vimentin was found in damaged epithelial cells with mesenchymal features. Conclusion: We demonstrated possible evidence to understand the role of SMP30 as a new pivotal factor and the possibility of decreased SMP30 as a potential indicator of EMT at the end stage of CKD. [ABSTRACT FROM AUTHOR]

Background: The senescence marker protein 30 (SMP30) is a calcium-binding protein whose expression decreases with age, and is closely associated with hepatocellular carcinoma (HCC) development. The primary goal of this study was to examine the mechanistic effect of SMP30 on HCC migration and invasion. Methods: Bioinformatic and immunohistochemical approaches were used to examine the expression of SMP30 in HCC tissues and its relationship to patient survival. We investigated the effects of SMP30 expression on HCC cell proliferation, migration, invasion, and cell cycle dynamics. cDNA microarray technology was used to determine the gene expression profile of SK-Hep-1 cells following recombinant SMP30 overexpression to identify genes downstream of SMP30 that regulate HCC cell migration and invasion. We identified SMP30 interacting proteins by affinity purification-mass spectrometry (AP-MS) and co-immunoprecipitation/western blotting (COIP-WB). Results: SMP30 expression was lower in HCC tissues compared with normal liver tissues, and its expression positively correlated with overall survival in HCC patients. Additionally, SMP30 overexpression effectively blocked the migratory and invasive properties of SK-Hep-1 cells, but did not affect either proliferation rates or cell cycle. cDNA microarray results confirmed that many of the differentially expressed genes identified are involved in the process of epithelial-mesenchymal transition (EMT). AP-MS and COIP-WB experiments confirmed that Rho-associated protein kinase 1 (ROCK1) interacts with SMP30 in SK-Hep-1 cells, and ROCK1 is known to intimately regulate the EMT process. Conclusion: SMP30 inhibits HCC metastasis by influencing the expression of EMT-related proteins after interacting with ROCK1.

Background: The senescence marker protein 30 (SMP30) is a calcium-binding protein whose expression decreases with age, and is closely associated with hepatocellular carcinoma (HCC) development. The primary goal of this study was to examine the mechanistic effect of SMP30 on HCC migration and invasion., Methods: Bioinformatic and immunohistochemical approaches were used to examine the expression of SMP30 in HCC tissues and its relationship to patient survival. We investigated the effects of SMP30 expression on HCC cell proliferation, migration, invasion, and cell cycle dynamics. cDNA microarray technology was used to determine the gene expression profile of SK-Hep-1 cells following recombinant SMP30 overexpression to identify genes downstream of SMP30 that regulate HCC cell migration and invasion. We identified SMP30 interacting proteins by affinity purification-mass spectrometry (AP-MS) and co-immunoprecipitation/western blotting (COIP-WB)., Results: SMP30 expression was lower in HCC tissues compared with normal liver tissues, and its expression positively correlated with overall survival in HCC patients. Additionally, SMP30 overexpression effectively blocked the migratory and invasive properties of SK-Hep-1 cells, but did not affect either proliferation rates or cell cycle. cDNA microarray results confirmed that many of the differentially expressed genes identified are involved in the process of epithelial-mesenchymal transition (EMT). AP-MS and COIP-WB experiments confirmed that Rho-associated protein kinase 1 (ROCK1) interacts with SMP30 in SK-Hep-1 cells, and ROCK1 is known to intimately regulate the EMT process., Conclusion: SMP30 inhibits HCC metastasis by influencing the expression of EMT-related proteins after interacting with ROCK1., Competing Interests: The authors declare no conflict of interest., (© 2024 The Author(s). Published by IMR Press.)

Background/aim: Chronic kidney disease (CKD) is one of the most common causes of mortality in wild non-domestic felidae. The molecular mechanism regulating renal fibrosis in nephropathy is not fully understood especially in the felidae. This study aimed to elucidate senescence marker protein 30 (SMP30) expression patterns and its relationship with epithelial-mesenchymal transition (EMT) by immunostaining in two necropsied Siberian tigers (Panthera tigris altaica) with CKD., Materials and Methods: Two kidney samples from male Siberian tigers were fixed and tissue sections were stained for histopathological assay., Results: In CKD, renal tubular epithelial cells lost their tubular structures surrounded by severe interstitial fibrosis and were detached from the basement membrane. These damaged cells resembled the morphology of mesenchymal cells and showed much lower SMP30 expression compared with intact tubular epithelial cells. These cells also expressed vimentin, which is specifically expressed by mesenchymal cells, and through double staining, it was observed that vimentin was expressed in the tubular epithelial cells where SMP30 was not expressed. In addition, double-positive expression of pan-cytokeratin (pan-CK) and vimentin was found in damaged epithelial cells with mesenchymal features., Conclusion: We demonstrated possible evidence to understand the role of SMP30 as a new pivotal factor and the possibility of decreased SMP30 as a potential indicator of EMT at the end stage of CKD., (Copyright © 2024, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.)