A Galectin-9-Driven CD11chigh Decidual Macrophage Subset Suppresses Uterine Vascular Remodeling in Preeclampsia.

BACKGROUND: Preeclampsia is a serious disease of pregnancy that lacks early diagnosis methods or effective treatment, except delivery. Dysregulated uterine immune cells and spiral arteries are implicated in preeclampsia, but the mechanistic link remains unclear. METHODS: Single-cell RNA sequencing and spatial transcriptomics were used to identify immune cell subsets associated with preeclampsia. Cell-based studies and animal models including conditional knockout mice and a new preeclampsia mouse model induced by recombinant mouse galectin-9 were applied to validate the pathogenic role of a CD11chigh subpopulation of decidual macrophages (dMφ) and to determine its underlying regulatory mechanisms in preeclampsia. A retrospective preeclampsia cohort study was performed to determine the value of circulating galectin-9 in predicting preeclampsia. RESULTS: We discovered a distinct CD11chigh dMφ subset that inhibits spiral artery remodeling in preeclampsia. The proinflammatory CD11chigh dMφ exhibits perivascular enrichment in the decidua from patients with preeclampsia. We also showed that trophoblast-derived galectin-9 activates CD11chigh dMφ by means of CD44 binding to suppress spiral artery remodeling. In 3 independent preeclampsia mouse models, placental and plasma galectin-9 levels were elevated. Galectin-9 administration in mice induces preeclampsia-like phenotypes with increased CD11chigh dMφ and defective spiral arteries, whereas galectin-9 blockade or macrophage-specific CD44 deletion prevents such phenotypes. In pregnant women, increased circulating galectin-9 levels in the first trimester and at 16 to 20 gestational weeks can predict subsequent preeclampsia onset. CONCLUSIONS: These findings highlight a key role of a distinct perivascular inflammatory CD11chigh dMφ subpopulation in the pathogenesis of preeclampsia. CD11chigh dMφ activated by increased galectin-9 from trophoblasts suppresses uterine spiral artery remodeling, contributing to preeclampsia. Increased circulating galectin-9 may be a biomarker for preeclampsia prediction and intervention.

Strategy and technique for surgical treatment of Ebstein's anomaly.

BACKGROUND: Ebstein's anomaly (EA) is a rare and complex congenital heart anomaly, and the effect of surgical treatment is not ideal. This study aims to introduce our experience in management strategies, surgical techniques, and operative indications for patients with Ebstein's anomaly. METHODS: A retrospective study of 258 operations was performed in 253 patients by the same cardiac surgeon in The First Hospital of Tsinghua University between March 2004 and January 2020. 32 patients had previously received cardiac surgery in other hospitals. The clinical data including diagnosis, operative indications, techniques, pathological changes, and survival rates were collected and analyzed. RESULTS: Anatomical correction was performed in 203 (78.7%) operations, 1½ ventricle repair in 38 (14.7%) operations, tricuspid valve repair only in four operations (1.6%), tricuspid valve replacement in ten (3.9%), total cavopulmonary connection (TCPC) in two (0.8%), and Glenn operation in one operation (0.4%). Reoperation was performed in five patients (2.0%) during hospitalization. Among them, tricuspid valve replacement was performed in one patient, 1½ ventricle repair in two patients, and tricuspid valve annulus reinforcement in two patients. Five patients died with an early mortality rate of 2.0%. Complete atrioventricular conduction block was complicated in one patient (0.4%). A total of 244 patients was followed up (four in the 253 patients lost) with a duration of 3.0-168.0 (87.6 ± 38.4) months. Cardiac function of 244 patients improved significantly with mean New York Heart Association (NYHA) functional class recovery from 3.5 to 1.1. The mean grade of tricuspid valve regurgitation improved from 3.6 to 1.5. Three late deaths (1.2%) occurred. The survival rates at five and ten years after surgery were 98.6% and 98.2%, respectively. Reoperation was performed in five patients (2.0%) during the follow-up period. CONCLUSION: Based on our management strategies and operative principles and techniques, anatomical correction of EA is capable of achieving excellent long-term results, and low rates of TCPC, 1½ ventricle repair and valvular replacement.

Surgical strategies and outcomes for myocardial bridges coexisting with other cardiac conditions.

BACKGROUND: Myocardial bridges are congenital coronary artery anomalies. There are still many controversies surrounding surgical treatment strategies for myocardial bridges combined with other heart disorders. The purpose of this study was to evaluate the surgical treatment strategies and outcomes in patients with these conditions. METHODS: Between March 2004 and October 2021, our institution witnessed 77 patients diagnosed with myocardial bridging who underwent surgical intervention. According to the myocardial bridge and combined heart disorder, four groups were identified: 1. isolated LAD supra-arterial myotomy group, 2. LAD CABG and(or not) myotomy  group, 3. LAD supra-arterial myotomy and grafting of other branches group, and 4. LAD supra-arterial myotomy and other cardiac surgery group. The perioperative outcomes, symptoms, life quality, mortality, and major adverse cardiac events (MACEs) were analyzed. RESULTS: There were no deaths during hospitalization and no rethoractomy for postoperative bleeding or major adverse cardiac events (MACEs). The follow-up period ranged from 2 months to 199.2 months (55.61 ± 10.21) months, the 10-year cumulative survival rates for the four groups of patients were 95.0%, 100%, 100% and 74.1%, and the 10-year freedom rates from the MACEs were 83.9%, 92.0%, 87.5% and 76.2%, respectively. CONCLUSIONS: Supra-arterial myotomy is preferred in patients with isolated myocardial bridge, and acceptable results can be achieved by choosing supra-arterial myotomy in combination with CABG or other cardiac surgery simultaneously for patients with myocardial bridges and other heart disorders.

Golgi α-mannosidases regulate cell surface N-glycan type and ectodomain shedding of the transmembrane protease corin.

Corin is a transmembrane protease that activates natriuretic peptides on the cell membrane. Reduced cell surface targeting or increased ectodomain shedding disrupts cell membrane homeostasis of corin, thereby impairing its cell surface expression and enzyme activity. N-glycans are essential in corin ectodomain shedding. Lack of N-glycans promotes corin ectodomain shedding in the juxtamembrane and frizzled-1 domains. The nascent N-glycans, transferred onto the polypeptide of corin, undergo multistep N-glycan processing in the endoplasmic reticulum and Golgi. It remains unclear how trimming by Golgi α-mannosidases, the critical N-glycan processing steps in N-glycan maturation, may regulate corin biosynthesis. In this study, we examined the effects of kifunensine and swainsonine, the inhibitors for α-mannosidases I and II, on corin expression and function. Western analysis of corin proteins in cell lysates and conditioned media from the inhibitor-treated corin-stable HEK293 cells and AC16 cells showed that both α-mannosidases I and II were required to maintain complex N-glycans on cell surface corin and protect corin from ectodomain shedding in the juxtamembrane and frizzled-1 domains. Cell viability analysis revealed that inhibition of α-mannosidase I or II sensitized cardiomyocytes to hydrogen peroxide-induced injury via regulating corin. Moreover, either one of the two coding genes was sufficient to perform Golgi α-mannosidase I trimming of N-glycans on corin. Similarly, this sufficiency was observed in Golgi α-mannosidase II-coding genes. Inhibition of ectodomain shedding restored corin zymogen activation from kifunensine- or swainsonine-induced reduction. Together, our results show the important roles of Golgi α-mannosidases in maintaining cell membrane homeostasis and biological activities of corin.

Natriuretic Peptide Signaling in Uterine Biology and Preeclampsia.

Endometrial decidualization is a uterine process essential for spiral artery remodeling, embryo implantation, and trophoblast invasion. Defects in endometrial decidualization and spiral artery remodeling are important contributing factors in preeclampsia, a major disorder in pregnancy. Atrial natriuretic peptide (ANP) is a cardiac hormone that regulates blood volume and pressure. ANP is also generated in non-cardiac tissues, such as the uterus and placenta. In recent human genome-wide association studies, multiple loci with genes involved in natriuretic peptide signaling are associated with gestational hypertension and preeclampsia. In cellular experiments and mouse models, uterine ANP has been shown to stimulate endometrial decidualization, increase TNF-related apoptosis-inducing ligand expression and secretion, and enhance apoptosis in arterial smooth muscle cells and endothelial cells. In placental trophoblasts, ANP stimulates adenosine 5'-monophosphate-activated protein kinase and the mammalian target of rapamycin complex 1 signaling, leading to autophagy inhibition and protein kinase N3 upregulation, thereby increasing trophoblast invasiveness. ANP deficiency impairs endometrial decidualization and spiral artery remodeling, causing a preeclampsia-like phenotype in mice. These findings indicate the importance of natriuretic peptide signaling in pregnancy. This review discusses the role of ANP in uterine biology and potential implications of impaired ANP signaling in preeclampsia.

Surgical intervention of myocardial bridge combined coronary artery disease: could a combination of supra-arterial myotomy and CABG be a better option?

BACKGROUND: The treatment of coronary artery disease combined with severe atherosclerotic stenosis proximal to a left anterior descending artery myocardial bridge (LAD-MB) is still controversial. This study aimed to analyze the outcomes of surgical intervention in patients with severe atherosclerotic stenosis proximal to a LAD-MB. METHODS: We retrospectively reviewed all patients with coronary artery disease combined with severe atherosclerotic stenosis proximal to the LAD-MB. The enrolled criteria were systolic compression of LAD more than or equal to 50% and atherosclerotic stenosis proximal to the LAD-MB more than or equal to 70%. All patients suffered from anginal symptoms refractory to medical therapy. All patients received supra-arterial myotomy and coronary artery bypass grafting (CABG) procedures. Clinical characteristics, intraoperative findings, and postoperative outcomes were evaluated. RESULTS: Between 2004 and 2021, sixteen patients underwent supra-arterial myotomy and CABG procedure. The compression and length of LAD-MB were 63 ± 17.9% and 25.9 ± 16.3 mm, respectively. Of the 16 patients, one patient had a LAD-MB and proximal coronary stenosis, and 15 patients had LAD-MBs and multivessel lesions. All patients survived and recovered uneventfully without in-hospital mortality or severe complications. The median transfusion amount of red blood cells in the operation was 2 units, and no patients required unplanned reoperation for bleeding. The average length of intensive care unit stay was 2.74 days. Fifteen patients were followed up for 6-146.1 months (mean 45.3 ± 42.9 months). One patient had a recurrence of angina pectoris one year after surgery, and 14 patients had no symptoms of myocardial ischemia during the follow-up period. Significant improvement in symptoms and quality of life using the Seattle Angina Questionnaire assessment was observed in all five categories after surgery (p < 0.01). CONCLUSIONS: Based on the results, supra-arterial myotomy and concomitant bypass surgery may be a better option for the treatment of LAD-MB combined with severe proximal stenosis.

Spatial position is a key determinant of N-glycan functionality of the scavenger receptor cysteine-rich domain of human hepsin.

The scavenger receptor cysteine-rich (SRCR) domain is a key constituent in diverse proteins. N-glycosylation is important in protein expression and function. In the SRCR domain of different proteins, N-glycosylation sites and functionality vary substantially. In this study, we examined the importance of N-glycosylation site positions in the SRCR domain of hepsin, a type II transmembrane serine protease involved in many pathophysiological processes. We analysed hepsin mutants with alternative N-glycosylation sites in the SRCR and protease domains using three-dimensional modelling, site-directed mutagenesis, HepG2 cell expression, immunostaining, and western blotting. We found that the N-glycan function in the SRCR domain in promoting hepsin expression and activation on the cell surface cannot be replaced by alternatively created N-glycans in the protease domain. Within the SRCR domain, the presence of an N-glycan in a confined surface area was essential for calnexin-assisted protein folding, endoplasmic reticulum (ER) exiting, and zymogen activation of hepsin on the cell surface. Hepsin mutants with alternative N-glycosylation sites on the opposite side of the SRCR domain were trapped by ER chaperones, resulting in the activation of the unfolded protein response in HepG2 cells. These results indicate that the spatial N-glycan positioning in the SRCR domain is a key determinant in the interaction with calnexin and subsequent cell surface expression of hepsin. These findings may help to understand the conservation and functionality of N-glycosylation sites in the SRCR domains of different proteins.

Transmembrane serine protease TMPRSS2 implicated in SARS-CoV-2 infection is autoactivated intracellularly and requires N-glycosylation for regulation.

Transmembrane protease serine 2 (TMPRSS2) is a membrane-bound protease expressed in many human epithelial tissues, including the airway and lung. TMPRSS2-mediated cleavage of viral spike protein is a key mechanism in severe acute respiratory syndrome coronavirus 2 activation and host cell entry. To date, the cellular mechanisms that regulate TMPRSS2 activity and cell surface expression are not fully characterized. In this study, we examined two major post-translational events, zymogen activation and N-glycosylation, in human TMPRSS2. In experiments with human embryonic kidney 293, bronchial epithelial 16HBE, and lung alveolar epithelial A549 cells, we found that TMPRSS2 was activated via intracellular autocatalysis and that this process was blocked in the presence of hepatocyte growth factor activator inhibitors 1 and 2. By glycosidase digestion and site-directed mutagenesis, we showed that human TMPRSS2 was N-glycosylated. N-glycosylation at an evolutionarily conserved site in the scavenger receptor cysteine-rich domain was required for calnexin-assisted protein folding in the endoplasmic reticulum and subsequent intracellular trafficking, zymogen activation, and cell surface expression. Moreover, we showed that TMPRSS2 cleaved severe acute respiratory syndrome coronavirus 2 spike protein intracellularly in human embryonic kidney 293 cells. These results provide new insights into the cellular mechanism in regulating TMPRSS2 biosynthesis and function. Our findings should help to understand the role of TMPRSS2 in major respiratory viral diseases.

Coronary artery bypass grafting in patients with malignancy: a single-institute case series of eight patients.

BACKGROUND: The surgical strategy among patients with malignancy and coronary artery disease (CAD) remains controversial. In this study, we present the experiences of coronary artery bypass grafting (CABG) in patients with malignancy and analyzed the treatment outcomes. METHODS: From January 2011 to October 2021, eight patients combined with coronary artery disease and malignancy, six of them with three-vessel disease and two with anterior descending branch lesions on coronary angiography. The age ranged from 54 to 73 years (61.8 ± 7.7years). Four patients underwent CABG and staging for surgical oncology, and 2 patients underwent CABG and surgical oncology simultaneously. Four patients underwent CABG procedure with cardiopulmonary bypass (on-pump CABG), and the other patients underwent the procedure without cardiopulmonary bypass (off-pump CABG). All patients were followed up for 3 to 96 months (40.4 ± 31.5 months) postoperatively. RESULTS: The mean number of grafts was 2.6 ± 1.1, there was no in-hospital death, postoperative myocardial infarction, and stroke. Among the eight patients, one patient received chemotherapy and radiation before bypass surgery, which occurred postoperatively pulmonary infection, and the rest of 7 patients had no major adverse cardiovascular events during follow-up periods. CONCLUSION: Based on the results of the present study, simultaneous or staged CABG and oncologic surgery according to the TNM stage of the tumor and cardiac assessment is an effective treatment for patients with severe CAD combined with malignancy.

Left ventricle reverse remodeling in chronic aortic regurgitation patients with dilated ventricle after aortic valve replacement.

BACKGROUND: Aortic valve replacement (AVR) for chronic aortic regurgitation (AR) with a severe dilated left ventricle and dysfunction leads to left ventricle remodeling. But there are rarely reports on the left ventricle reverse remodeling (LVRR) after AVR. This study aimed to investigate the LVRR and outcomes in chronic AR patients with severe dilated left ventricle and dysfunction after AVR. METHODS: We retrospectively analyzed the clinical datum of chronic aortic regurgitation patients who underwent isolated AVR. The LVRR was defined as an increase in left ventricular ejection fraction (LVEF) at least 10 points or a follow-up LVEF ≥ 50%, and a decrease in the indexed left ventricular end-diastolic diameter of at least 10%, or an indexed left ventricular end-diastolic diameter ≤ 33 mm/m2. The changes in echocardiographic parameters after AVR, survival analysis, the predictors of major adverse cardiac events (MACE), the association between LVRR and MACE were analyzed. RESULTS: Sixty-nine patients with severe dilated left ventricle and dysfunction underwent isolated AVR. LV remodeling in 54 patients and no LV remodeling in 15 patients at 6-12 months follow-up. The preoperative left ventricular dimensions and volumes were larger, and the EF was lower in the LV no remodeling group than those in the LV remodeling group (all p < 0.05). The adverse LVRR was the predictor for MACE at follow-up. The mean follow-up period was 47.29 months (range 6 to 173 months). The rate of freedom from MACE was 94.44% at 5 years and 92.59% at 10 years in the remodeling group, 60% at 5 years, and 46.67% at 10 years in the no remodeling group. CONCLUSIONS: The left ventricle remodeling after AVR was the important predictor for MACE. LV no remodeling may not be associated with benefits from AVR for chronic aortic regurgitation patients with severe dilated LV and dysfunction.

CD320 expression and apical membrane targeting in renal and intestinal epithelial cells.

Vitamin B12 is an essential nutrient acquired via dietary intake. Receptor-mediated endocytosis is a key mechanism in vitamin B12 absorption, cellular uptake, and reabsorption. CD320 is a type I transmembrane protein responsible for cellular uptake of vitamin B12 in peripheral tissues. In this study, we examined segmental distribution and cellular expression of CD320 in mouse kidneys and intestines. We show that CD320 is expressed on the luminal surface in the small intestine and in proximal tubules in the kidney, suggesting that, in addition to its role in vitamin B12 uptake in peripheral tissues, CD320 may participate in vitamin B12 absorption in the small intestine and reabsorption in the kidney. Moreover, we show that an amino acid motif, DSSDE, in the second low-density lipoprotein receptor class A domain of CD320 is a key apical membrane targeting signal in both renal and intestinal epithelial cells. Mutations or deletion of this motif abolish the specific apical membrane expression of CD320 in polarized Madin-Darby canine kidney cells and human colon cancer-derived Caco-2 cells. In short-hairpin RNA-based gene knockdown experiments, we show that the apical membrane targeting of CD320 is mediated by a Rab11a-dependent mechanism. These results extend our knowledge regarding the cell biology of CD320 and its role in vitamin B12 metabolism.

Hepsin regulates TGFß signaling via fibronectin proteolysis.

Transforming growth factor-beta (TGFß) is a multifunctional cytokine with a well-established role in mammary gland development and both oncogenic and tumor-suppressive functions. The extracellular matrix (ECM) indirectly regulates TGFß activity by acting as a storage compartment of latent-TGFß, but how TGFß is released from the ECM via proteolytic mechanisms remains largely unknown. In this study, we demonstrate that hepsin, a type II transmembrane protease overexpressed in 70% of breast tumors, promotes canonical TGFß signaling through the release of latent-TGFß from the ECM storage compartment. Mammary glands in hepsin CRISPR knockout mice showed reduced TGFß signaling and increased epithelial branching, accompanied by increased levels of fibronectin and latent-TGFß1, while overexpression of hepsin in mammary tumors increased TGFß signaling. Cell-free and cell-based experiments showed that hepsin is capable of direct proteolytic cleavage of fibronectin but not latent-TGFß and, importantly, that the ability of hepsin to activate TGFß signaling is dependent on fibronectin. Altogether, this study demonstrates a role for hepsin as a regulator of the TGFß pathway in the mammary gland via a novel mechanism involving proteolytic downmodulation of fibronectin.

Atrial natriuretic peptide promotes uterine decidualization and a TRAIL-dependent mechanism in spiral artery remodeling.

Atrial natriuretic peptide (ANP) is an important hormone in cardiovascular biology. It is activated by the protease corin. In pregnancy, ANP and corin promote uterine spiral artery remodeling, but the underlying mechanism remains unknown. Here we report an ANP function in uterine decidualization and TNF-related apoptosis-inducing ligand-dependent (TRAIL-dependent) death in spiral arterial smooth muscle cells (SMCs) and endothelial cells (ECs). In ANP- or corin-deficient mice, uterine decidualization markers and TRAIL expression were decreased, whereas in cultured human endometrial stromal cells (HESCs), ANP increased decidualization and TRAIL expression. In uterine spiral arteries from pregnant wild-type mice, SMC and EC loss occurred sequentially before trophoblast invasion. In culture, TRAIL from decidualized HESCs induced apoptosis in uterine SMCs, but not in ECs with low TRAIL receptor expression. Subsequently, cyclophilin B was identified from apoptotic SMCs that upregulated endothelial TRAIL receptor and caused apoptosis in ECs. These results indicate that ANP promotes decidualization and TRAIL expression in endometrial stromal cells, contributing to sequential events in remodeling of spiral arteries, including SMC death and cyclophilin B release, which in turn induces TRAIL receptor expression and apoptosis in ECs.

Surgical intervention and outcome for treatment of myocardial bridging refractory to medication.

INTRODUCTION: Myocardial bridging (MB) is a common and usually benign inborn coronary abnormality that may lead to anginal symptoms, acute coronary syndrome, arrhythmias, and rarely sudden cardiac death. MB are most commonly localized in the middle segment of the left anterior descending coronary artery (LAD). The treatment of LAD-MB is still challenging. Our objective was to assess the short- and long-term results of surgical procedures in patients with LAD-MB who had chest pain refractory to medical therapy. METHODS: Between March 2005 and January 2020, 26 patients (19 males and 7 females; mean 55.8 ± 12.4 years) with MB underwent surgery. All MB was located in the mid-segment of the LAD with a mean length of 4.2 ± 1.7 cm. Coronary angiography before surgery demonstrated LAD-MB with systolic compression more than or equal to 70% in all patients. RESULTS: Twenty-five patients underwent myotomy and one patient underwent coronary artery bypass grafting (CABG). All patients survived and recovered uneventfully. Neither hospital or late death nor major complications occurred. Follow-up time was 3-173 months (mean 55.7 months). Follow-up of coronary angiography or computed tomography scan performed in 16 patients demonstrated restoration of coronary blood flow and myocardial perfusion without significant residual compression of the artery. All patients were symptom-free and are currently in NYHA Class I. CONCLUSION: The symptomatic LAD-MB patients who are refractory to medication should actively undergo the surgical intervention such as myotomy and CABG to eliminate the clinical symptoms and achieve satisfactory results by follow-up findings. Myotomy is a preferred procedure because of its safety and satisfactory results.

Novel Trilaminar Polymeric Antiadhesion Membrane Prevents Postoperative Pericardial Adhesion.

BACKGROUND: Postoperative pericardial adhesion formation is a prominent cause of morbidity and death in cardiovascular surgery, but there is still no ideal prevention method, especially in redo cases. This study investigated a novel antiadhesion trilaminar polymeric film compared with the Gore PRECLUDE Pericardial Membrane (W. L. Gore & Associates, Flagstaff, AZ) and a negative control. METHODS: Our novel trilaminar membrane is composed of polyvinyl alcohol (PVA) and carboxymethylcellulose (CMC). An established pericardial adhesion rabbit model was used to test the property of the membrane. After sternotomy, a portion of pericardium was resected and the epicardium was abraded. Rabbits (n = 24) were randomly assigned to 3 groups: control group, no trilaminar membrane or expanded polytetrafluoroethylene (ePTFE); ePTFE group, or trilaminar membrane group (PVA-CMC group). Evaluation of adhesion formation was performed by resternotomy 4 weeks after the operation. RESULTS: The degree of tissue adhesion surrounding the heart in the PVA-CMC group was significantly less than in the control (P < .01) and ePTFE (P < .05) groups. The inflammation score in group PVA-CMC was significantly lower than that in the ePTFE (P < .01) and control (P < .01) groups. The fibrosis score was significantly lower in group PVA-CMC than that in the ePTFE (P < .05) and control (P < .01) groups. CONCLUSIONS: The novel trilaminar membrane effectively reduced postoperative pericardial adhesions. The placement of the trilaminar membrane at the time of sternal closure provides a novel combination to minimize the extent and severity of pericardial adhesions while providing a physical barrier between the sternum and the cardiac structures.

Hepsin: a multifunctional transmembrane serine protease in pathobiology.

Cell membrane-bound serine proteases are important in the maintenance of physiological homeostasis. Hepsin is a type II transmembrane serine protease highly expressed in the liver. Recent studies indicate that hepsin activates prohepatocyte growth factor in the liver to enhance Met signaling, thereby regulating glucose, lipid, and protein metabolism. In addition, hepsin functions in nonhepatic tissues, including the adipose tissue, kidney, and inner ear, to regulate adipocyte differentiation, urinary protein processing, and auditory function, respectively. In mouse models, hepsin deficiency lowers blood glucose, lipid, and protein levels, impairs uromodulin assembly in renal epithelial cells, and causes hearing loss. Elevated hepsin expression has also been found in many cancers. As a type II transmembrane protease, cell surface expression and zymogen activation are essential for hepsin activity. In this review, we discuss the current knowledge regarding hepsin biosynthesis, activation, and functions in pathobiology.

A conserved LDL-receptor motif regulates corin and CD320 membrane targeting in polarized renal epithelial cells.

Selective protein distribution on distinct plasma membranes is important for epithelial cell function. To date, how proteins are directed to specific epithelial cell surface is not fully understood. Here we report a conserved DSSDE motif in LDL-receptor (LDLR) modules of corin (a transmembrane serine protease) and CD320 (a receptor for vitamin B12 uptake), which regulates apical membrane targeting in renal epithelial cells. Altering this motif prevents specific apical corin and CD320 expression in polarized Madin-Darby canine kidney (MDCK) cells. Mechanistic studies indicate that this DSSDE motif participates in a Rab11a-dependent mechanism that specifies apical sorting. In MDCK cells, inhibition of Rab11a, but not Rab11b, expression leads to corin and CD320 expression on both apical and basolateral membranes. Together, our results reveal a novel molecular recognition mechanism that regulates LDLR module-containing proteins in their specific apical expression in polarized renal epithelial cells.

Intracellular autoactivation of TMPRSS11A, an airway epithelial transmembrane serine protease.

Type II transmembrane serine proteases (TTSPs) are a group of enzymes participating in diverse biological processes. Some members of the TTSP family are implicated in viral infection. TMPRSS11A is a TTSP expressed on the surface of airway epithelial cells, which has been shown to cleave and activate spike proteins of the severe acute respiratory syndrome (SARS) and the Middle East respiratory syndrome coronaviruses (CoVs). In this study, we examined the mechanism underlying the activation cleavage of TMPRSS11A that converts the one-chain zymogen to a two-chain enzyme. By expression in human embryonic kidney 293, esophageal EC9706, and lung epithelial A549 and 16HBE cells, Western blotting, and site-directed mutagenesis, we found that the activation cleavage of human TMPRSS11A was mediated by autocatalysis. Moreover, we found that TMPRSS11A activation cleavage occurred before the protein reached the cell surface, as indicated by studies with trypsin digestion to remove cell surface proteins, treatment with cell organelle-disturbing agents to block intracellular protein trafficking, and analysis of a soluble form of TMPRSS11A without the transmembrane domain. We also showed that TMPRSS11A was able to cleave the SARS-CoV-2 spike protein. These results reveal an intracellular autocleavage mechanism in TMPRSS11A zymogen activation, which differs from the extracellular zymogen activation reported in other TTSPs. These findings provide new insights into the diverse mechanisms in regulating TTSP activation.

In vitro study on electrospun lecithin-based poly (L-lactic acid) scaffolds and their biocompatibility.

Lecithin is a natural mixture of phospholipids and neutral lipids, and plays a vital role in the maintenance of cell-membrane integrity for all of the basic biological processes.This study aimed to evaluate the adhesion and growth of bone marrow-derived mesenchymal stem cells (BMSCs) on the three-dimensional (3D) electrospun lecithin-containing poly (L-lactic acid) (PLLA) fibrous scaffolds. In this study, PLLA and 0-15% of lecithin (wt.%) were dissolved in methylene chloride and thoroughly mixed to fabricate fibrous scaffolds by electrospinning technique. Surface morphology and hydrophilic properties of the scaffolds were characterized by scanning electron microscopy (SEM) and water uptake, respectively. The cytocompatibility of electrospun scaffolds and their potential cytotoxic effects on cells were studied over 8 days by seeding rat BMSCs on the scaffolds of PLLA containing 0-15% lecithin. Then cell proliferation was tested by CCK-8 assay and cell injury was assessed by measuring the release of intracellular lactate dehydrogenase (LDH). And cell morphology was studied by Toluidine blue staining, propidium iodide staining and SEM. The hydrophilicity was dramatically increased with an increase of lecithin content in the modified PLLA/lecithin scaffolds, as determined by rate of water uptake. In addition, the results obtained from in vitro assays suggested that the electrospun PLLA/5%lecithin scaffolds could possess the optimal hydrophilicity, higher adhesion and proliferation of BMSCs than PLLA, and they also did not lead to any significant toxic effects to cells. Moreover, BMSCs on PLLA/5%lecithin scaffolds tended to maintain their phenotypic shape and integrate with the microfibers to form a 3D cellular network, indicating a favorable interaction between this electrospun lecithin-based scaffolds and cells. The study demonstrated that the electrospun PLLA/lecithin fibrous scaffolds showed much better hydrophilicity and biocompatibility by introducing lecithin into the polymer in comparison with pure PLLA, which would offer a promising strategy for constructing tissue engnineered blood vessel.

Fatal course of clofazimine-induced pulmonary crystal deposition in a patient with Melkersson-Rosenthal syndrome.

A wide variety of drugs and substances have the potential to damage the respiratory system by different mechanisms. Clofazimine is an anti-leprosy drug that is normally only prescribed for a few years. It has a very long half-life, and crystalline deposition of the drug in various tissues has been documented. But up to now, no fatalities due to pulmonary damage have been described. We report the case of a patient who took clofazimine for almost 27 years as off-label treatment for Melkersson-Rosenthal syndrome. He suffered from progressive dyspnea, productive cough, and occasional hemoptysis. X-ray and CT of the thoracic organs revealed extensive multilocular, compact, tumor-like infiltrates with central necrosis in both lungs. Pulmonary function tests showed restrictive impairment and manifest hypoxemia. Histology of lung biopsies revealed intense interstitial accumulation of histiocytes and marked deposition of crystalline foreign material. The patient died from progressive respiratory failure. Autopsy revealed crystalline deposition and a histiocytic reaction in many other parenchymal organs. Conclusion: Pulmonary parenchymal deposition of drug crystals is a rare mechanism of drug-induced pulmonary diseases. Long-standing, off-label use of clofazimine may cause severe destruction of the lungs and can be fatal.