Augmented ERAD (ER-associated degradation) activity in chondrocytes is necessary for cartilage development and maintenance.

Chondrocytes secrete massive extracellular matrix (ECM) molecules that are produced, folded, and modified in the endoplasmic reticulum (ER). Thus, the ER-associated degradation (ERAD) complex-which removes misfolded and unfolded proteins to maintain proteostasis in the ER- plays an indispensable role in building and maintaining cartilage. Here, we examined the necessity of the ERAD complex in chondrocytes for cartilage formation and maintenance. We show that ERAD gene expression is exponentially increased during chondrogenesis, and disruption of ERAD function causes severe chondrodysplasia in developing embryos and loss of adult articular cartilage. ERAD complex malfunction also causes abnormal accumulation of cartilage ECM molecules and subsequent chondrodysplasia. ERAD gene expression is decreased in damaged cartilage from patients with osteoarthritis (OA), and disruption of ERAD function in articular cartilage leads to cartilage destruction in a mouse OA model.

Effect of a confirmatory testing algorithm on early acute HIV diagnosis in Korea.

ABSTRACT: The 17 Provincial Institutes of Health and Environment (PIHEs) in Korea use HIV antibody, antigen, and Western blot assays for confirmatory testing of HIV infection. The Korea Disease Control and Prevention Agency (KDCA) has further included p24 antigen neutralization and nucleic acid tests (NATs) since 2015. Our study aimed to investigate the effect of this new testing algorithm on the confirmation rate of HIV infection.Annual changes, from 2012 through 2017, in positive or indeterminate HIV confirmatory results were compared for the two algorithms between the PIHEs and the KDCA. Fiebig stages and Western blot p31 band were used to identify the diagnostic proportions of acute or early chronic HIV for the two algorithms.The number of positive cases in the samples requested from PIHEs for reconfirmation by the KDCA has steadily increased from 10.3% in 2014 to 33.3% in 2017. However, the number of indeterminate cases dropped sharply, from 71.9% in 2014 to 14.0% in 2017. The results for the p31 reactive band were 27.4% and 88.4% for the KDCA and PIHEs, respectively. Of positive cases reported by the KDCA, 22.9% were in the early acute stage and Fiebig stages I to II.The new testing algorithm has improved the diagnosis of HIV infections in the early acute stage. Early confirmatory diagnosis can prevent secondary transmission of HIV and provide early treatment opportunities for people living with HIV infection.

Extracellular matrixes and neuroinflammation.

The extracellular matrix is a critical component of every human tissue. ECM not only functions as a structural component but also regulates a variety of cellular processes such as cell migration, differentiation, proliferation, and cell death. In addition, current studies suggest that ECM is critical for the pathophysiology of various human diseases. ECM is composed of diverse components including several proteins and polysaccharide chains such as chondroitin sulfate, heparan sulfate, and hyaluronic acid. Each component of ECM exerts its own functions in cellular and pathophysiological processes. One of the interesting recent findings is that ECM is involved in inflammatory responses in various human tissues. In this review, we summarized the known functions of ECM in neuroinflammation after acute injury and chronic inflammatory diseases of the central nerve systems. [BMB Reports 2020; 53(10): 491-499].

Alpha-tocopherol exerts protective function against the mucotoxicity of particulate matter in amphibian and human goblet cells.

Exposure to particulate matter (PM) in ambient air is known to increase the risk of cardiovascular disorders and mortality. The cytotoxicity of PM is mainly due to the abnormal increase of reactive oxygen species (ROS), which damage cellular components such as DNA, RNA, and proteins. The correlation between PM exposure and human disorders, including mortality, is based on long-term exposure. In this study we have investigated acute responses of mucus-secreting goblet cells upon exposure to PM derived from a heavy diesel engine. To this end, we employed the mucociliary epithelium of amphibian embryos and human Calu-3 cells to examine PM mucotoxicity. Our data suggest that acute exposure to PM significantly impairs mucus secretion and results in the accumulation of mucus vesicles in the cytoplasm of goblet cells. RNA-seq analysis revealed that acute responses to PM exposure significantly altered gene expression patterns; however, known regulators of mucus production and the secretory pathway were not significantly altered. Interestingly, pretreatment with α-tocopherol nearly recovered the hyposecretion of mucus from both amphibian and human goblet cells. We believe this study demonstrates the mucotoxicity of PM and the protective function of α-tocopherol on mucotoxicity caused by acute PM exposure from heavy diesel engines.

Simple Method To Characterize the Ciliary Proteome of Multiciliated Cells.

Motile cilia of multiciliated epithelial cells have important roles in animal development and cell homeostasis. Although several studies have identified and reported proteins localized in this complex organelle and the related immotile primary cilia from various cell types, it is still challenging to isolate high quantities of ciliary proteins for proteomic analysis. In this study, African clawed frog (Xenopus laevis) embryos, which have many multiciliated cells in the epidermis, were treated with a simple ionic buffer to identify 1009 proteins conserved across vertebrates; these proteins were putatively localized in motile cilia. Using two ciliary proteome databases, we confirmed that previously validated cilia-associated proteins are highly enriched in our ciliary proteome. Proteins localized at the transition zone and Ellis-van Creveld zone, which are distinct regions at the base of cilia, near the junction with the apical cell surface, were isolated using our method. Among the newly identified ciliary proteins, we report that KRT17 may have an unrecognized function in motile cilia. Hence, the method developed in this study would be useful for understanding the ciliary proteome.

ITGBL1 modulates integrin activity to promote cartilage formation and protect against arthritis.

Developing and mature chondrocytes constantly interact with and remodel the surrounding extracellular matrix (ECM). Recent research indicates that integrin-ECM interaction is differentially regulated during cartilage formation (chondrogenesis). Integrin signaling is also a key source of the catabolic reactions responsible for joint destruction in both rheumatoid arthritis and osteoarthritis. However, we do not understand how chondrocytes dynamically regulate integrin signaling in such an ECM-rich environment. Here, we found that developing chondrocytes express integrin-ß-like 1 (Itgbl1) at specific stages, inhibiting integrin signaling and promoting chondrogenesis. Unlike cytosolic integrin inhibitors, ITGBL1 is secreted and physically interacts with integrins to down-regulate activity. We observed that Itgbl1 expression was strongly reduced in the damaged articular cartilage of patients with osteoarthritis (OA). Ectopic expression of Itgbl1 protected joint cartilage against OA development in the destabilization of the medial meniscus-induced OA mouse model. Our results reveal ITGBL1 signaling as an underlying mechanism of protection against destructive cartilage disorders and suggest the potential therapeutic utility of targeting ITGBL1 to modulate integrin signaling in human disease.

Xenopus: An alternative model system for identifying muco-active agents.

The airway epithelium in human plays a central role as the first line of defense against environmental contaminants. Most respiratory diseases such as chronic obstructive pulmonary disease (COPD), asthma, and respiratory infections, disturb normal muco-ciliary functions by stimulating the hypersecretion of mucus. Several muco-active agents have been used to treat hypersecretion symptoms in patients. Current muco-active reagents control mucus secretion by modulating either airway inflammation, cholinergic parasympathetic nerve activities or by reducing the viscosity by cleaving crosslinking in mucin and digesting DNAs in mucus. However, none of the current medication regulates mucus secretion by directly targeting airway goblet cells. The major hurdle for screening potential muco-active agents that directly affect the goblet cells, is the unavailability of in vivo model systems suitable for high-throughput screening. In this study, we developed a high-throughput in vivo model system for identifying muco-active reagents using Xenopus laevis embryos. We tested mucus secretion under various conditions and developed a screening strategy to identify potential muco-regulators. Using this novel screening technique, we identified narasin as a potential muco-regulator. Narasin treatment of developing Xenopus embryos significantly reduced mucus secretion. Furthermore, the human lung epithelial cell line, Calu-3, responded similarly to narasin treatment, validating our technique for discovering muco-active reagents.

Blockade of cannabinoid 1 receptor improves glucose responsiveness in pancreatic beta cells.

Cannabinoid 1 receptors (CB1Rs) are expressed in peripheral tissues, including islets of Langerhans, where their function(s) is under scrutiny. Using mouse ß-cell lines, human islets and CB1R-null (CB1R-/- ) mice, we have now investigated the role of CB1Rs in modulating ß-cell function and glucose responsiveness. Synthetic CB1R agonists diminished GLP-1-mediated cAMP accumulation and insulin secretion as well as glucose-stimulated insulin secretion in mouse ß-cell lines and human islets. In addition, silencing CB1R in mouse ß cells resulted in an increased expression of pro-insulin, glucokinase (GCK) and glucose transporter 2 (GLUT2), but this increase was lost in ß cells lacking insulin receptor. Furthermore, CB1R-/- mice had increased pro-insulin, GCK and GLUT2 expression in ß cells. Our results suggest that CB1R signalling in pancreatic islets may be harnessed to improve ß-cell glucose responsiveness and preserve their function. Thus, our findings further support that blocking peripheral CB1Rs would be beneficial to ß-cell function in type 2 diabetes.

Crystal structure of SEL1L: Insight into the roles of SLR motifs in ERAD pathway.

Terminally misfolded proteins are selectively recognized and cleared by the endoplasmic reticulum-associated degradation (ERAD) pathway. SEL1L, a component of the ERAD machinery, plays an important role in selecting and transporting ERAD substrates for degradation. We have determined the crystal structure of the mouse SEL1L central domain comprising five Sel1-Like Repeats (SLR motifs 5 to 9; hereafter called SEL1L(cent)). Strikingly, SEL1L(cent) forms a homodimer with two-fold symmetry in a head-to-tail manner. Particularly, the SLR motif 9 plays an important role in dimer formation by adopting a domain-swapped structure and providing an extensive dimeric interface. We identified that the full-length SEL1L forms a self-oligomer through the SEL1L(cent) domain in mammalian cells. Furthermore, we discovered that the SLR-C, comprising SLR motifs 10 and 11, of SEL1L directly interacts with the N-terminus luminal loops of HRD1. Therefore, we propose that certain SLR motifs of SEL1L play a unique role in membrane bound ERAD machinery.

A thioredoxin fold protein Sh3bgr regulates Enah and is necessary for proper sarcomere formation.

The sh3bgr (SH3 domain binding glutamate-rich) gene encodes a small protein containing a thioredoxin-like fold, SH3 binding domain, and glutamate-rich domain. Originally, it was suggested that increased expression of Sh3bgr may cause the cardiac phenotypes in Down's syndrome. However, it was recently reported that the overexpression of Sh3bgr did not cause any disease phenotypes in mice. In this study, we have discovered that Sh3bgr is critical for sarcomere formation in striated muscle tissues and also for heart development. Sh3bgr is strongly expressed in the developing somites and heart in Xenopus. Morpholino mediated-knockdown of sh3bgr caused severe malformation of heart tissue and disrupted segmentation of the somites. Further analysis revealed that Sh3bgr specifically localized to the Z-line in mature sarcomeres and that knockdown of Sh3bgr completely disrupted sarcomere formation in the somites. Moreover, overexpression of Sh3bgr resulted in abnormally discontinues thick firmaments in the somitic sarcomeres. We suggest that Sh3bgr does its function at least partly by regulating localization of Enah for the sarcomere formation. In addition, we provide the data supporting Sh3bgr is also necessary for proper heart development in part by affecting the Enah protein level.

Immune status and epidemiological characteristics of human immunodeficiency virus seroconverters in Korea, 1999-2009.

OBJECTIVES: The detection of HIV seroconverters increased annually since HIV antigen/antibody testing kits have been available widely in South Korea. This study aimed to identify the epidemiological characteristics of HIV seroconverters and their immune level at HIV diagnosis. METHOD: We analyzed the epidemiological and immunological characteristics of 341 HIV seroconverters among 6,008 HIV-diagnosed individuals from 1999 and 2009. The analysis of immune level and epidemiological factors of HIV seroconverters was conducted by using chi-square test on SAS version 9.1. RESULTS: The seroconverters among newly-identified HIV cases each year increased from 0.5% in 1999 to over 5% or in 2009. The sex ratio of seroconverters was 18:1 (male:female), and 33% were in their 30s, and 28% were in their 20s. Reasons for HIV testing were involvement in voluntary test due to risky behaviors (43%), and health check-up (36%). Discovery of HIV infection occurred primarily in hospitals (84%). Among seroconverters, 55 percent had a CD4 T-cell count of more than 350/µl. CONCLUSION: Korean HIV seroconverters tended to be younger at diagnosis, diagnosed during a voluntary test, and their CD4+ T-cell counts at HIV diagnosis were higher than those of non-seroconverters aall HIV-infected individuals. This study of HIV seroconverters will be important foundational in future studies on HIV incidence, disease progress, and survival rate.