Cathepsin Research Articles

Usefulness of Cathepsin S to Predict Risk for Obstructive Sleep Apnea among Patients with Type 2 Diabetes

Background Obstructive sleep apnea (OSA) was highly prevalent in patients with type 2 diabetes (T2D). Cathepsin S (CTSS), a cysteine protease, is involved in the inflammatory activity in T2D and hypoxia conditions. The aim of the study was to evaluate whether CTSS could be involved in the inflammatory reaction of OSA in patients with T2D. Methods We included 158 participants in this study matched for age, gender, and body mass index in 4 groups (control, non-OSA&T2D, OSA&non-T2D, and OSA&T2D). After overnight polysomnography, we collected the clinical data including anthropometrical characteristics, blood pressure, and fasting blood samples in the morning. Plasma CTSS concentration was evaluated using the human Magnetic Luminex Assay. Results Compared with the control group, both the non-OSA&T2D group and the OSA&non-T2D group showed higher CTSS levels. Plasma CTSS expression was significantly increased in subjects with OSA&T2D compared to subjects with non-OSA&T2D. The OSA&T2D group had higher CTSS levels than the OSA&non-T2D group, but there were no statistically significant differences. Plasma CTSS levels showed significant correlation with the apnea-hypopnea index (AHI) (r = 0.559, P < 0.001) and plasma fasting blood glucose (r = 0.427, P < 0.001). After adjusting confounding factors, plasma CTSS levels were independently associated with the AHI (Beta: 0.386, 95% confidence intervals (CI): 21.988 to 57.781; P < 0.001). Furthermore, we confirmed the higher pinpoint accuracy of plasma CTSS in the diagnosis of OSA (area under the curve: 0.868). Conclusions Plasma CTSS expression was significantly elevated in the OSA&T2D group and was independently associated with the AHI; it could be a biomarker with a positive diagnostic value on diagnosing OSA among patients with T2D.

BAF60a Deficiency in Vascular Smooth Muscle Cells Prevents Abdominal Aortic Aneurysm by Reducing Inflammation and Extracellular Matrix Degradation.

Currently, there are no approved drugs for abdominal aortic aneurysm (AAA) treatment, likely due to limited understanding of the primary molecular mechanisms underlying AAA development and progression. BAF60a-a unique subunit of the SWI/SNF (switch/sucrose nonfermentable) chromatin remodeling complex-is a novel regulator of metabolic homeostasis, yet little is known about its function in the vasculature and pathogenesis of AAA. In this study, we sought to investigate the role and underlying mechanisms of vascular smooth muscle cell (VSMC)-specific BAF60a in AAA formation. Approach and Results: BAF60a is upregulated in human and experimental murine AAA lesions. In vivo studies revealed that VSMC-specific knockout of BAF60a protected mice from both Ang II (angiotensin II)-induced and elastase-induced AAA formation with significant suppression of vascular inflammation, monocyte infiltration, and elastin fragmentation. Through RNA sequencing and pathway analysis, we found that the expression of inflammatory response genes in cultured human aortic smooth muscle cells was significantly downregulated by small interfering RNA-mediated BAF60a knockdown while upregulated upon adenovirus-mediated BAF60a overexpression. BAF60a regulates VSMC inflammation by recruiting BRG1 (Brahma-related gene-1)-a catalytic subunit of the SWI/SNF complex-to the promoter region of NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells) target genes. Furthermore, loss of BAF60a in VSMCs prevented the upregulation of the proteolytic enzyme cysteine protease CTSS (cathepsin S), thus ameliorating ECM (extracellular matrix) degradation within the vascular wall in AAA. Our study demonstrated that BAF60a is required to recruit the SWI/SNF complex to facilitate the epigenetic regulation of VSMC inflammation, which may serve as a potential therapeutic target in preventing and treating AAA.

Nicotine Modulates CTSS (Cathepsin S) Synthesis and Secretion Through Regulating the Autophagy-Lysosomal Machinery in Atherosclerosis.

Increased CTSS (cathepsin S) has been reported to play a critical role in atherosclerosis progression. Both CTSS synthesis and secretion are essential for exerting its functions. However, the underlying mechanisms contributing to CTSS synthesis and secretion in atherosclerosis remain unclear. Approach and Results: In this study, we showed that nicotine activated autophagy and upregulated CTSS expression in vascular smooth muscle cells and in atherosclerotic plaques. Western blotting and immunofluorescent staining showed that nicotine inhibited the mTORC1 (mammalian target of rapamycin complex 1) activity, promoted the nuclear translocation of TFEB (transcription factor EB), and upregulated the expression of CTSS. Chromatin immunoprecipitation-qualificative polymerase chain reaction, electrophoretic mobility shift assay, and luciferase reporter assay further demonstrated that TFEB directly bound to the CTSS promoter. mTORC1 inhibition by nicotine or rapamycin promoted lysosomal exocytosis and CTSS secretion. Live cell assays and IP-MS (immunoprecipitation-mass spectrometry) identified that the interactions involving Rab10 (Rab10, member RAS oncogene family) and mTORC1 control CTSS secretion. Nicotine promoted vascular smooth muscle cell migration by upregulating CTSS, and CTSS inhibition suppressed nicotine-induced atherosclerosis in vivo. We concluded that nicotine mediates CTSS synthesis and secretion through regulating the autophagy-lysosomal machinery, which offers a potential therapeutic target for atherosclerosis treatment.

Insecticidal Activity of Artemisia vulgaris Essential Oil and Transcriptome Analysis of Tribolium castaneum in Response to Oil Exposure

Red flour beetle (Tribolium castaneum) is one of the most destructive pests of stored cereals worldwide. The essential oil (EO) of Artemisia vulgaris (mugwort) is known to be a strong toxicant that inhibits the growth, development, and reproduction of T. castaneum. However, the molecular mechanisms underlying the toxic effects of A. vulgaris EO on T. castaneum remain unclear. Here, two detoxifying enzymes, carboxylesterase (CarEs) and cytochrome oxidase P450 (CYPs), were dramatically increased in red flour beetle larvae when they were exposed to A. vulgaris EO. Further, 758 genes were differentially expressed between EO treated and control samples. Based on Gene Ontology (GO) analysis, numerous differentially expressed genes (DEGs) were enriched for terms related to the regulation of biological processes, response to stimulus, and antigen processing and presentation. Our results indicated that A. vulgaris EO disturbed the antioxidant activity in larvae and partially inhibited serine protease (SP), cathepsin (CAT), and lipase signaling pathways, thus disrupting larval development and reproduction as well as down-regulating the stress response. Moreover, these DEGs showed that A. vulgaris indirectly affected the development and reproduction of beetles by inducing the expression of genes encoding copper-zinc-superoxide dismutase (CuZnSOD), heme peroxidase (HPX), antioxidant enzymes, and transcription factors. Moreover, the majority of DEGs were mapped to the drug metabolism pathway in the Kyoto Encyclopedia of Genes and Genomes (KEGG) database. Notably, the following genes were detected: 6 odorant binding proteins (OBPs), 5 chemosensory proteins (CSPs), 14 CYPs, 3 esterases (ESTs), 5 glutathione S-transferases (GSTs), 6 UDP-glucuronosyltransferases (UGTs), and 2 multidrug resistance proteins (MRPs), of which 8 CYPs, 2 ESTs, 2 GSTs, and 3 UGTs were up-regulated dramatically after exposure to A. vulgaris EO. The residual DEGs were significantly down-regulated in EO exposed larvae, implying that partial compensation of metabolism detoxification existed in treated beetles. Furthermore, A. vulgaris EO induced overexpression of OBP/CYP, and RNAi against these genes significantly increased mortality of larvae exposed to EO, providing further evidence for the involvement of OBP/CYP in EO metabolic detoxification in T. castaneum. Our results provide an overview of the transcriptomic changes in T. castaneum in response to A. vulgaris EO.

Pharmacological Inhibition of Cathepsin S Suppresses Abdominal Aortic Aneurysm in Mice

Evidence suggests that cathepsin S (CTSS), a potent mammalian elastase, participates in abdominal aortic aneurysm (AAA) formation. This study examines the hypothesis that pharmacological inhibition of CTSS with an α-ketoamide based compound 6r might suppress AAA in mice.

Decreased serum cathepsin S levels in patients with systemic sclerosis-associated interstitial lung disease.

Cathepsin S (CTSS) is a lysosomal proteolytic enzyme regulating intracellular and extracellular biological activities, including immunity/inflammation and remodeling of vasculature and extracellular matrix, which are the three cardinal pathological events associated with systemic sclerosis (SSc). To elucidate the potential role of CTSS in the development of SSc, we investigated the clinical correlation of serum CTSS levels. Because serum CTSS levels were inversely correlated with estimated glomerular filtration rate (eGFR) in SSc patients with renal dysfunction (eGFR, <60min/mL per 1.73m2 ), SSc patients with normal renal function (eGFR, ≥60min/mL per 1.73m2 ) were analyzed. Serum CTSS levels were significantly decreased in diffuse cutaneous SSc patients compared with limited cutaneous SSc patients and healthy controls. Among vascular and fibrotic clinical manifestations, Raynaud's phenomenon and interstitial lung disease (ILD) were relevant to a significant decrease in serum CTSS levels. Importantly, serum CTSS levels negatively correlated with serum levels of Krebs von den Lungen-6 and surfactant protein D in total SSc patients, while not correlating with modified Rodnan total skin thickness score and the percentage of predicted diffusion lung capacity for carbon monoxide and showing a positive trend with the percentage of predicted vital capacity. These results suggest a potential contribution of decreased CTSS expression to the development of ILD in patients with SSc.

Cathepsin S Protease Mediates T-Cell Response in Non-Hodgkin Lymphoma

Cathepsin S (CTSS) aberrant activation or overexpression promotes non-Hodgkin lymphoma development.

Inverse relationship between autophagy and CTSK is related to bovine embryo quality.

Improving the quality and the developmental competence of in vitro produced (IVP) embryos is an indispensable goal for assisted reproductive technology. Autophagy is a major protective mechanism for intracellular degradation of unnecessary cytoplasmic components. Autophagy ends by the fusion between autophagic vacuoles and lysosomes, allowing the degradation of the cargo by lysosomal enzymes, especially the cathepsins (CTSs). However, it is still unclear how autophagy and cathepsin K (CTSK) relate to embryo development. This study evaluated (1.) the activities of autophagy and CTSK in relation to bovine embryo quality and (2.) the effect of autophagy induction and/or CTSK inhibition on preimplantation embryo development and quality. We show here that good-quality embryos exhibited a greater autophagic activity and less CTSK activity compared to poor-quality embryos. Blastomeres of an individual embryo may vary in their quality. Good quality blastomeres showed an increased autophagic activity and decreased CTSK activity compared to poor-quality blastomeres within the same embryo at different developmental stages. Importantly, induction of autophagy and/or inhibition of CTSK improved the developmental rate (increased blastocyst and hatching rates) and the quality (increased total cell number and decreased the percentage of apoptotic cells) of IVP bovine embryos. These results demonstrate a promising approach to selectively isolate good-quality embryos and improve the efficiency of IVEP of cattle embryos.

Novel nanobiosensor based protease biomarkers for early detection and prognosis of pancreatic cancers.

e16787 Background: Identifying novel biomarkers for early detection and prognosis would allow for improved survival outcomes in pancreatic cancer (PC). Arginase has been implicated in tumorigenesis through its production of L-ornithine. Its aberrant expression provides metabolites for tumor proliferation. Methods: In our study, 47 patients with metastatic PC, 29 with localized PC, and 50 healthy controls were enrolled at KU Cancer Center from 2000 to 2019. Overall survival (OS) was retrospectively assessed and correlated to serum protease levels at diagnosis. Protease expression was analyzed using nanobiosensors, which utilized fluorescent nanoparticles read by Spectra Scan to measure arginase, matrix metalloproteinases (MMPs), urokinase plasminogen activator (uPA), neutrophil elastase (NE), cathepsin B (CTSB), and cathepsin E (CTSE). Survival analysis was completed by Kaplan-Meier method. Results: Protease expression in localized vs. healthy cases was significant for (CTSB [p = 1.33E-10], MMP1 [p = 6.66E-21], MMP3 [p = 1.39E-13], MMP9 [p = 2.77E-08], NE [p = 0.00015], uPA [p = 2.55E-11]) and borderline significant for arginase [p = 0.05082]. Protease expression in metastatic vs. healthy cases was significant for (CTSB [p = 7.06E-06], MMP1 [p = 3.08E-12], MMP3 [p = 2.51E-14], MMP9 [p = 1.73E-08], NE [p = 0.01430], uPA [p = 0.00024]). Further, metastatic vs. localized cancer was significant for (arginase [p = 0.00034], CTSB [p = 0.00584], MMP1 [p = 2.03E-13], NE [p = 2.29E-10], and uPA [p = 6.58E-07]. Characteristics of 21 localized PC patients are illustrated in Table. Median OS was 25m in localized PC with low Arginase expression (mean &lt; 215) vs 17.5m in high Arginase expression (p = 0.0477). Conclusions: Expression pattern of CTSB, MMP, NE, arginase, uPA proteases showed statistical significant difference for detection of localized PC vs metastatic PC vs healthy cases. In localized PC, lower arginase expression showed a statistically significant improvement in survival vs high arginase expression. Arginase expression was borderline significant for detection of localized PC vs healthy controls, likely due to small sample. Arginase as a potential biomarker for early detection and prognosis of PC warrants validation in a larger cohort. [Table: see text]

Cathepsin S Alterations Induce a Tumor-Promoting Immune Microenvironment in Follicular Lymphoma

Tumor cells orchestrate their microenvironment. Here, we provide biochemical, structural, functional, and clinical evidence that Cathepsin S (CTSS) alterations induce a tumor-promoting immune microenvironment in follicular lymphoma (FL). We found CTSS mutations at Y132 in 6% of FL (19/305). Another 13% (37/286) had CTSS amplification, which was associated with higher CTSS expression. CTSS Y132 mutations lead to accelerated autocatalytic conversion from an enzymatically inactive profrom to active CTSS and increased substrate cleavage, including CD74, which regulates major histocompatibility complex class II (MHC class II)-restricted antigen presentation. Lymphoma cells with hyperactive CTSS more efficiently activated antigen-specific CD4+ Tcells invitro. Tumors with hyperactive CTSS showed increased CD4+ Tcell infiltration and proinflammatory cytokine perturbation in a mouse model and in human FLs. In mice, this CTSS-induced immune microenvironment promoted tumor growth. Clinically, patients with CTSS-hyperactive FL had better treatment outcomes with standard immunochemotherapies, indicating that these immunosuppressive regimens target both the lymphoma cells and the tumor-promoting immune microenvironment.

Cathepsin S Regulates Antigen Processing and T Cell Activity in Non-Hodgkin Lymphoma

Genomic alterations in cancer cells can influence the immune system to favor tumor growth. In non-Hodgkin lymphoma, physiological interactions between B cells and the germinal center microenvironment are coopted to sustain cancer cell proliferation. We found that follicular lymphoma patients harbor a recurrent hotspot mutation targeting tyrosine 132 (Y132D) in cathepsin S (CTSS) that enhances protein activity. CTSS regulates antigen processing and CD4+ and CD8+ Tcell-mediated immune responses. Loss of CTSS activity reduces lymphoma growth by limiting communication with CD4+ T follicular helper cells while inducing antigen diversification and activation of CD8+ Tcells. Overall, our results suggest that CTSS inhibition has non-redundant therapeutic potential to enhance anti-tumor immune responses in indolent and aggressive lymphomas.

Prognostic role and therapeutic susceptibility of cathepsin in various types of solid tumor and leukemia: A systematic review.

Cathepsins (CTSs) are multifunctional proteins that can play prominent roles in cancer progression and metastasis. In this systematic review, we compared the prognosis of CTS subtypes overexpression in leukemia and solid tumors, and investigated the effect of different factors on CTS prognosis. We systematically searched published articles indexed in PubMed, Scopus, Cochrane library, ISI Web of Science, and EmBase databases from February 2000 until January 2020. Among the selected leukemia and solid tumors studies, overexpression of CTS subtypes in newly diagnosed and treated patients were with poor prognosis in 43 studies (79.6%) and with good prognosis in 9 studies (16.6%). However, there were 2 studies (3.8%) with either good or poor prognosis, depending on conditions and caner stage and host cell. The relation between CTS and human leukocyte antigen (HLA) in leukemia and solid tumors was mentioned in 7 studies (13%). Overexpression of CTS subtypes in all new case patients had contributed to the induction of poor prognosis. It seems that CTS subtypes, based on the type of cancer and its stage, the type of host cells, and the probable relation with HLA, breed good or poor prognosis in patients with cancer. Therefore, monitoring the overexpression of CTS subtypes and determining the effect of each of these factors on CTS prognosis could be helpful in predicting cancer prognosis both in newly diagnosed or under treatment patients. They could also be useful in finding ways for improving the efficiency of contemporary therapeutic strategies in various types of leukemia and solid tumors.

Catching Cathepsin X in the Act

Catching Cathepsin X in the Act

Genome-wide identification, expression signature and immune functional analysis of two cathepsin S (CTSS) genes in turbot (Scophthalmus maximus L.)

Cathepsins, a superfamily of hydrolytic enzymes produced and enclosed within lysosomes, play multiple roles at physiological and pathological states. Cathepsin S is a lysosomal cysteine endopeptidase of the papain family, and exerts critical roles in the regulation of MHC class II immune responses. In the present study, we captured two Cathepsin S genes in turbot (SmCTSS1 and SmCTSS2.1), characterized their expression patterns following V. anguillarum and S. iniae infections, and explored their binding ability and agglutination capability. Firstly, the SmCTSS1 contained a 990 bp ORF encoding 329 amino acids, while SmCTSS2.1 contained a 1,014 bp ORF encoding 337 amino acids. The phylogenetic analysis revealed that both genes showed the closest relationship to their counterparts of Japanese flounder (Paralichthys olivaceus). In addition, both genes were ubiquitously expressed in all examined healthy tissues, with the highest expression level observed in spleen and intestine, respectively, while the lowest expression level both observed in liver. Both SmCTSS1 and SmCTSS2.1 were significantly differentially expressed, and exhibited general down-regulations at most time points in skin and intestine after two bacterial infections. Finally, both rSmCTSS1 and rSmCTSS2.1 showed significant binding ability to three examined microbial ligands (LPS, PGN and LTA), and strong agglutination effect to different bacteria (E. tarda, S. agalactiae, S. aureus and V. anguillarum). Collectively, this study provided valuable data for understanding the roles of CTSS in the host defense against bacterial infections in turbot, and indicated the potential vital roles of CTSS in innate immune responses of teleost species.

Cathepsins in the Pathophysiology of Mucopolysaccharidoses: New Perspectives for Therapy.

Cathepsins (CTSs) are ubiquitously expressed proteases normally found in the endolysosomal compartment where they mediate protein degradation and turnover. However, CTSs are also found in the cytoplasm, nucleus, and extracellular matrix where they actively participate in cell signaling, protein processing, and trafficking through the plasma and nuclear membranes and between intracellular organelles. Dysregulation in CTS expression and/or activity disrupts cellular homeostasis, thus contributing to many human diseases, including inflammatory and cardiovascular diseases, neurodegenerative disorders, diabetes, obesity, cancer, kidney dysfunction, and others. This review aimed to highlight the involvement of CTSs in inherited lysosomal storage disorders, with a primary focus to the emerging evidence on the role of CTSs in the pathophysiology of Mucopolysaccharidoses (MPSs). These latter diseases are characterized by severe neurological, skeletal and cardiovascular phenotypes, and no effective cure exists to date. The advance in the knowledge of the molecular mechanisms underlying the activity of CTSs in MPSs may open a new challenge for the development of novel therapeutic approaches for the cure of such intractable diseases.

MiR-491-3p is down-regulated in postmenopausal osteoporosis and affects growth, differentiation and apoptosis of hFOB1.19 cells through targeting CTSS.

Postmenopausal osteoporosis (PMO) is a common disease related to aging, which has been paid increasing attention in recent years because of its serious complications. MiR-491-3p was found to play a crucial roles in several diseases. However, the role of miR-491-3p in PMO has yet not been studied. Our research intends to explore the impact of miR-491-3p on PMO in the in vitro model. The expression patterns of miR-491-3p and cathepsin S (CTSS) in patients with PMO were acquired from the GEO database. The human osteoblast cell hFOB1.19 was used to detect the function of miR-491-3p and CTSS in PMO. The viability and apoptosis of hFOB1.19 cells were measured by cell counting kit 8 and flow cytometry assays. The apoptosis and differentiation related proteins were analyzed by western blotting. The relationship between miR-491-3p and CTSS was predicted by appropriate software and affirmed by luciferase assay. MiR-491-3p expression was lower in patients with PMO. The up-regulation of miR-491-3p in hFOB1.19 cells increased their viability and differentiation and inhibited their apoptosis. CTSS, which was highly expressed in patients with PMO, was confirmed as a direct target of miR-491-3p and was found to be inversely modulated by miR-491-3p. The rescue assays showed that overexpression of CTSS suppressed the promoting effects of miR-491-3p mimic on the proliferation and differentiation of hFOB1.19 cells, and repressed the inhibitory effects of miR-491-3p mimic on apoptosis of hFOB1.19 cells. The results of our study showed that miR-491-3p could ameliorate biological characteristics of hFOB1.19 cells by reducing CTSS expression suggesting that miR-491-3p/CTSS might be a potential biomarker for the diagnosis and treatment of PMO.

Tissue-Specific Factors Differentially Regulate the Expression of Antigen-Processing Enzymes During Dendritic Cell Ontogeny.

Dendritic cells (DCs) form a collection of antigen-presenting cells (APCs) that are distributed throughout the body. Conventional DCs (cDCs), which include the cDC1 and cDC2 subsets, and plasmacytoid DCs (pDCs) constitute the two major ontogenically distinct DC populations. The pDCs complete their differentiation in the bone marrow (BM), whereas the cDC subsets derive from pre-committed BM precursors, the pre-cDC, that seed lymphoid and non-lymphoid tissues where they further differentiate into mature cDC1 and cDC2. Within different tissues, cDCs express distinct phenotype and function. Notably, cDCs in the thymus are exquisitely efficient at processing and presenting antigens in the class II pathway, whereas in the spleen they do so only upon maturation induced by danger signals. To appraise this functional heterogeneity, we examined the regulation of the expression of distinct antigen-processing enzymes during DC ontogeny. We analyzed the expression of cathepsin S (CTSS), cathepsin L (CTSL), and thymus-specific serine protease (TSSP), three major antigen-processing enzymes regulating class II presentation in cDC, by DC BM precursors and immature and mature cDCs from the spleen and thymus. We found that pre-cDCs in the BM express relatively high levels of these different proteases. Then, their expression is modulated in a tissue-specific and subset-specific manner with immature and mature thymic cDCs expressing overall higher levels than immature splenic cDCs. On the other hand, the TSSP expression level is selectively down-regulated in spleen pDCs, whereas CTSS and CTSL are both increased in thymic and splenic pDCs. Hence, tissue-specific factors program the expression levels of these different proteases during DC differentiation, thus conferring tissue-specific function to the different DC subsets.

Paired associative stimulation improves recovery of neurological function after cerebral infarction through modulating neuronal autophagy

Objective To investigate the effect of paired associative stimulation (PAS) on the recovery of neurological function after cerebral infarction, and to explore whether any such effect is associated with autophagy in the ischemic penumbra. Methods Sixty adult male Sprague-Dawley rats were randomly divided into a sham operation group (n=20) and an experimental group (n=40). The rats of the experimental group underwent 90 minutes of right middle cerebral artery occlusion (MCAO), while the sham group received a sham operation. The experimental group was subsequently divided into a model group (n=20) and a PAS group (n=20). The PAS group received 14 days of paired associative stimulation (PAS) beginning 24 hours after the operation. Neurological dysfunction was evaluated with a modified neurological severity scale (mNSS) and the elevated body swing test (EBST) on the 1st, 7th and 14th day after the MCAO. The rats were then euthanized and the expression of LC3Ⅰ, LC3Ⅱ, Beclin1, and Cathepsin B in the ischemic penumbra were detected using Western blotting, while the distribution of LC3 in neurons was detected using double immunofluorescent staining. Results Compared with the sham group, the average mNSS scores and EBST values of the model and PAS groups were both higher on the 7th and 14th day after the MCAO, with those of the PAS group significantly lower than those of the model group on those days. The average mNSS score on the 14th day was significantly lower than on the 7th day. Compared with the sham group, the average LC3Ⅱ/Ⅰ values, Beclin 1 and Cathepsin B levels of both the model group and the PAS group were significantly higher on the 7th and 14th day after the MCAO, with the LC3Ⅱ/Ⅰ values of the PAS group significantly lower than those of the model group at both time points. The PAS group also had significantly lower Beclin1 and Cathepsin B levels on day 14. On the 7th and 14th days after the MCAO, the average number of LC3-positive cells and the ratio of LC3-positive neurons to total neurons in the model and PAS groups were significantly greater than the those of sham group, with the PAS group′s values significantly lower than those of the model group at each time point. Conclusion PAS can significantly promote neurological recovery after stroke. The beneficial effects may involve inhibition of neuronal autophagy in the ischemic penumbra. Key words: Cerebral infarction; Paired associative stimulation; Neuronal autophagy

Primary Alcohol-Activated Human and Mouse Hepatic Stellate Cells Share Similarities in Gene-Expression Profiles.

Alcoholic liver disease (ALD) is a leading cause of cirrhosis in the United States, which is characterized by extensive deposition of extracellular matrix proteins and formation of a fibrous scar. Hepatic stellate cells (HSCs) are the major source of collagen type 1 producing myofibroblasts in ALD fibrosis. However, the mechanism of alcohol‐induced activation of human and mouse HSCs is not fully understood. We compared the gene‐expression profiles of primary cultured human HSCs (hHSCs) isolated from patients with ALD (n = 3) or without underlying liver disease (n = 4) using RNA‐sequencing analysis. Furthermore, the gene‐expression profile of ALD hHSCs was compared with that of alcohol‐activated mHSCs (isolated from intragastric alcohol‐fed mice) or CCl4‐activated mouse HSCs (mHSCs). Comparative transcriptome analysis revealed that ALD hHSCs, in addition to alcohol‐activated and CCl4‐activated mHSCs, share the expression of common HSC activation (Col1a1 [collagen type I alpha 1 chain], Acta1 [actin alpha 1, skeletal muscle], PAI1 [plasminogen activator inhibitor‐1], TIMP1 [tissue inhibitor of metalloproteinase 1], and LOXL2 [lysyl oxidase homolog 2]), indicating that a common mechanism underlies the activation of human and mouse HSCs. Furthermore, alcohol‐activated mHSCs most closely recapitulate the gene‐expression profile of ALD hHSCs. We identified the genes that are similarly and uniquely up‐regulated in primary cultured alcohol‐activated hHSCs and freshly isolated mHSCs, which include CSF1R (macrophage colony‐stimulating factor 1 receptor), PLEK (pleckstrin), LAPTM5 (lysosmal‐associated transmembrane protein 5), CD74 (class I transactivator, the invariant chain), CD53, MMP9 (matrix metallopeptidase 9), CD14, CTSS (cathepsin S), TYROBP (TYRO protein tyrosine kinase‐binding protein), and ITGB2 (integrin beta‐2), and other genes (compared with CCl4‐activated mHSCs). Conclusion: We identified genes in alcohol‐activated mHSCs from intragastric alcohol‐fed mice that are largely consistent with the gene‐expression profile of primary cultured hHSCs from patients with ALD. These genes are unique to alcohol‐induced HSC activation in two species, and therefore may become targets or readout for antifibrotic therapy in experimental models of ALD.

Pharmacological Inhibition of Cathepsin S Suppresses Abdominal Aortic Aneurysm in Mice

Evidence suggests that cathepsin S (CTSS), a potent mammalian elastase, participates in abdominal aortic aneurysm (AAA) formation. This study examines the hypothesis that pharmacological inhibition of CTSS with an α-ketoamide based compound 6r might suppress AAA in mice. Experimental study of the CaCl2 induced AAA model in B6 mice and angiotensin II (AngII) infused AAA model in ApoE-/- mice. The effects of intraperitoneal administration of 6r (25mg/kg) and vehicle every three days since one day after AAA induction were evaluated at 28 days using CaCl2 induced (n=12 per group) and AngII infused (n=8 per group) models. Additionally, the effects of post-treatment with 6r and vehicle from seven days or 14 days after AAA induction were evaluated at 28 days using the CaCl2 induced model (n=6 per group). Aortic samples were harvested for histological and biochemical analyses, including cathepsin levels, Verhoeff Van Gieson staining, TUNEL assay, and immunostaining for macrophages. In the CaCl2 induced model, treatment with 6r suppressed aortic dilatation observed in vehicle treated controls (median: 0.58 vs. 0.92mm; p<.001), along with reduced CTSS and cathepsin K (CTSK) levels (both p<.001), preserved elastin integrity (p<.001), fewer medial apoptotic cells (p=.012) and less macrophage infiltration (p=.041). In the AngII infused model, the aortic diameter was smaller in 6r treated mice than in vehicle treated controls (median: 0.95 vs. 1.84mm; p=.047). The levels of CTSS (p<.001) and CTSK (p=.033) and the numbers of elastin breaks (p<.001), medial apoptotic cells (p<.001) and infiltrating macrophages (p=.030) were attenuated under 6r treatment. Finally, post-treatment with 6r from seven days (p=.046) or 14 days (p=.012) after AAA induction limited CaCl2 induced AAA. Pharmacological inhibition of CTSS by 6r suppresses AAA formation in mice. Also, post-treatment with 6r retards mouse AAA progression. These findings provide proof of concept validation for CTSS as a potential therapeutic target in AAA.