Loss of PBX1 function in Leydig cells causes testicular dysgenesis and male sterility.

Leydig cells are essential components of testicular interstitial tissue and serve as a primary source of androgen in males. A functional deficiency in Leydig cells often causes severe reproductive disorders; however, the transcriptional programs underlying the fate decisions and steroidogenesis of these cells have not been fully defined. In this study, we report that the homeodomain transcription factor PBX1 is a master regulator of Leydig cell differentiation and testosterone production in mice. PBX1 was highly expressed in Leydig cells and peritubular myoid cells in the adult testis. Conditional deletion of Pbx1 in Leydig cells caused spermatogenic defects and complete sterility. Histological examinations revealed that Pbx1 deletion impaired testicular structure and led to disorganization of the seminiferous tubules. Single-cell RNA-seq analysis revealed that loss of Pbx1 function affected the fate decisions of progenitor Leydig cells and altered the transcription of genes associated with testosterone synthesis in the adult testis. Pbx1 directly regulates the transcription of genes that play important roles in steroidogenesis (Prlr, Nr2f2 and Nedd4). Further analysis demonstrated that deletion of Pbx1 leads to a significant decrease in testosterone levels, accompanied by increases in pregnenolone, androstenedione and luteinizing hormone. Collectively, our data revealed that PBX1 is indispensable for maintaining Leydig cell function. These findings provide insights into testicular dysgenesis and the regulation of hormone secretion in Leydig cells.

Machine learning developed a CD8+ exhausted T cells signature for predicting prognosis, immune infiltration and drug sensitivity in ovarian cancer.

CD8+ exhausted T cells (CD8+ Tex) played a vital role in the progression and therapeutic response of cancer. However, few studies have fully clarified the characters of CD8+ Tex related genes in ovarian cancer (OC). The CD8+ Tex related prognostic signature (TRPS) was constructed with integrative machine learning procedure including 10 methods using TCGA, GSE14764, GSE26193, GSE26712, GSE63885 and GSE140082 dataset. Several immunotherapy benefits indicators, including Tumor Immune Dysfunction and Exclusion (TIDE) score, immunophenoscore (IPS), TMB score and tumor escape score, were used to explore performance of TRPS in predicting immunotherapy benefits of OC. The TRPS constructed by Enet (alpha = 0.3) method acted as an independent risk factor for OC and showed stable and powerful performance in predicting clinical outcome of patients. The C-index of the TRPS was higher than that of tumor grade, clinical stage, and many developed signatures. Low TRPS score indicated a higher level of CD8+ T cell, B cell, macrophage M1, and NK cells, representing a relative immunoactivated ecosystem in OC. OC patients with low risk score had a higher PD1&CTLA4 immunophenoscore, higher TMB score, lower TIDE score and lower tumor escape score, suggesting a better immunotherapy response. Moreover, higher TRPS score indicated a higher score of cancer-related hallmarks, including angiogenesis, EMT, hypoxia, glycolysis, and notch signaling. Vitro experiment showed that ARL6IP5 was downregulated in OC tissues and inhibited tumor cell proliferation. The current study constructed a novel TRPS for OC, which could serve as an indicator for predicting the prognosis, immune infiltration and immunotherapy benefits for OC patients.

Engineered extracellular vesicles: A new approach for targeted therapy of tumors and overcoming drug resistance.

Targeted delivery of anti-tumor drugs and overcoming drug resistance in malignant tumor cells remain significant clinical challenges. However, there are only few effective methods to address these issues. Extracellular vesicles (EVs), actively secreted by cells, play a crucial role in intercellular information transmission and cargo transportation. Recent studies have demonstrated that engineered EVs can serve as drug delivery carriers and showed promising application prospects. Nevertheless, there is an urgent need for further improvements in the isolation and purification of EVs, surface modification techniques, drug assembly processes, and precise recognition of tumor cells for targeted drug delivery purposes. In this review, we summarize the applications of engineered EVs in cancer treatment and overcoming drug resistance, and current challenges associated with engineered EVs are also discussed. This review aims to provide new insights and potential directions for utilizing engineered EVs as targeted delivery systems for anti-tumor drugs and overcoming drug resistance in the near future.

Molecular characterization and antimicrobial activity of NK-lysin in black scraper (Thamnaconus modestus).

NK-lysin (NKL) is a positively charged antimicrobial peptide with broad-spectrum bactericidal activities. In this study, the cDNA sequence of NKL (TmNKL) from black scraper (Thamnaconus modestus) was cloned, which encodes a predicted polypeptide of 150 amino acids that contains a surfactant protein B domain with three disulfide bonds. Phylogenetically, TmNKL was most closely related to its teleost counterpart from tiger puffer (Takifugu rubripes). Expression analysis demonstrated that TmNKL transcripts were constitutively expressed in all tested tissues, with the highest expression levels in the gills. Its expression was significantly upregulated in the gills, head kidney, and spleen after infection with Vibrio parahaemolyticus. A linear peptide (TmNKLP40L) and a disulfide-type peptide (TmNKLP40O) were further synthesized and results showed that disulfide bonds are not essential for bactericidal activities of TmNKL, and that both forms of TmNKL exhibited potent bactericidal activities against 4 gram- negative bacteria, including V. parahaemolyticus, V. alginolyticus, Edwardsiella tarda, and V. harveyi. Observed antimicrobial activities are likely due to the effects of TmNKLP40L and TmNKLP40O treatment on disrupting the integrity of both inner and outer membrane of V. parahaemolyticus, resulting in hydrolysis of bacterial genomic DNA. Damaged cell membranes and leakage of intracellular contents were further confirmed using scanning and transmission microscopy. Moreover, administration of 1.0 µg/g TmNKLP40L or TmNKLP40O significantly decreased bacterial load in tissues and thus, pronouncedly enhanced the survival of V. parahaemolyticus-infected fish. Overall, our results demonstrated that TmNKL is a potent innate effector and provides protective effects against bacterial infection.

Current understanding of the cGAS-STING signaling pathway: Structure, regulatory mechanisms, and related diseases.

The innate immune system protects the host from external pathogens and internal damage in various ways. The cGAS-STING signaling pathway, comprised of cyclic GMP-AMP synthase (cGAS), stimulator of interferon genes (STING), and downstream signaling adaptors, plays an essential role in protective immune defense against microbial DNA and internal damaged-associated DNA and is responsible for various immune-related diseases. After binding with DNA, cytosolic cGAS undergoes conformational change and DNA-linked liquid-liquid phase separation to produce 2'3'-cGAMP for the activation of endoplasmic reticulum (ER)-localized STING. However, further studies revealed that cGAS is predominantly expressed in the nucleus and strictly tethered to chromatin to prevent binding with nuclear DNA, and functions differently from cytosolic-localized cGAS. Detailed delineation of this pathway, including its structure, signaling, and regulatory mechanisms, is of great significance to fully understand the diversity of cGAS-STING activation and signaling and will be of benefit for the treatment of inflammatory diseases and cancer. Here, we review recent progress on the above-mentioned perspectives of the cGAS-STING signaling pathway and discuss new avenues for further study.

Molecular and functional characterization of a ladderlectin-like molecule from ayu (Plecoglossus altivelis).

Ladderlectin is a member of C-type lectins (CTLs) in teleost fish and involved in innate immune defense. In this study, ayu (Plecoglossus altivelis) ladderlecin-like (PaLL-like) sequence was cloned, which encodes a polypeptide of 172 amino acids that includes a signal peptide and characteristic C-type lectin-like domains (CTLDs). Phylogenetically, PaLL-like was most closely related to its teleost counterpart from shishamo smelt (Spirinchus lanceolatus). Expression analysis revealed a ubiquitous expression profile, with highest expression detected in liver and its expression was up-regulated following Vibiro anguillarum infection. Similar to canonical CTLs, PaLL-like exhibited carbohydrate-binidng capacities to a wide range of well-defined mono-/di-saccharides and likely confer PaLL-like the ability to agglutinate all tested bacterial, including three Gram-positive species (i.e., Listeria monocytogenes, Staphylococcus aureus and Streptococcus iniae) and eight Gram-negative species (i.e., Edwardsiella tarda, Aeromonas (A.) hydrophila, Escherichia coli, Vibrio (V.) harveyi, V. anguillarum, V. parahemolyticus, A. versoni and V. vulnificus), in a calcium-dependent manner. Further functional studies revealed that PaLL-like displayed immunomodulatory activities leading to enhanced bactericidal activity of serum, pathogen opsonization and macrophage activation with increased expression of pro-inflammatory cytokines (i.e., PaIL-1ß and PaTNF-α). Collectively, these immunomodulatory activities of PaLL-like suppressed proliferations of V. anguillarum in targeted tissued in vivo and likely contributed to the increased survival rate of infected-fish. Overall, our results demonstrated PaLL-like is a critical component of innate immunity and provides protective effects against bacterial infection.

A state-of-the-art review on LSD1 and its inhibitors in breast cancer: Molecular mechanisms and therapeutic significance.

Breast cancer (BC) is a kind of malignant cancer in women, and it has become the most diagnosed cancer worldwide since 2020. Histone methylation is a common biological epigenetic modification mediating varieties of physiological and pathological processes. Lysine-specific demethylase 1 (LSD1), a first identified histone demethylase, mediates the removal of methyl groups from histones H3K4me1/2 and H3K9me1/2 and plays a crucial role in varieties of cancer progression. It is also specifically amplified in breast cancer and contributes to BC tumorigenesis and drug resistance via both demethylase and non-demethylase manners. This review will provide insight into the overview structure of LSD1, summarize its action mechanisms in BC, describe the therapeutic potential of LSD1 inhibitors in BC, and prospect the current opportunities and challenges of targeting LSD1 for BC therapy.

Multimodal Neuroelectrophysiological Monitoring Combined with Robot-Assisted Placement of a Transiliac-Transsacral Screw for the Treatment of Transforaminal Sacral Fractures.

Objective: This study aimed to evaluate the safety and efficacy of the fixation of transforaminal sacral fractures using TiRobot-assisted transiliac-transsacral (TITS) screws under multimodal neuroelectrophysiological monitoring (MNM). Methods: From January 2019 to May 2021, 22 patients (17 male and 5 female patients) with transforaminal sacral fractures who were treated with closed reduction and placement of TiRobot-assisted TITS screws under MNM were retrospectively evaluated. The average age of the patients was 43.32 ± 11.40 years (range: 19-63). The patients received MNM, including somatosensory-evoked potentials (SEPs), motor-evoked potentials (MEPs), and electromyographic monitoring (EMG), prior to surgery, during closed reduction and the placement of the guidewire and TITS screw, and at the end of surgery. The operation was adjusted according to the MNM results. Results: Overall, 22 TITS screws were inserted in 22 patients, including 5 TITS screws in the S1 body and 17 TITS screws in the S2 body. The average time needed for screw placement was 27.95 ± 6.84 mins, and the average frequency of X-ray fluoroscopy exposures was 31.00 ± 5.56 for each patient. Anterior ring fixation was performed in 4 patients using an external fixator, in 5 patients using cannulated screws, and in 13 patients using reconstruction plates. The mean follow-up time was 14.46 ± 2.46 months (12-20 months). Tornetta and Matta radiographic outcomes were excellent in 10 patients, good in 9 patients, fair in 2 patients, and poor in 1 patient. The proportion of excellent and good ratings was 86.36%. At the final follow-up, the average Majeed score was 82.18 ± 14.52, with clinical outcomes that were excellent in 9 patients, good in 9 patients, fair in 1 patient, and poor in 3 patients. The proportion of excellent and good ratings was 82.82%. Preoperatively, the amplitude of the SEP on the injured side was lower than that on the contralateral side before reduction in 9 patients (>50%). In this study, no screw was mistakenly inserted into the sacral canal, and no surgical site infection occurred. Conclusion: MNM combined with TiRobot assistance can safely implant TITS screws and can effectively identify the neurological function of patients under anesthesia and reduce iatrogenic nerve injury.

[Effect of rat allogeneic BMSCs-Bio-Oss-bFGF compound on tooth extraction healing: a micro-CT study].

PRUPOSE: To investigate the effect of a compound of BMSCs-Bio-Oss-bFGF on microstructure of extraction sockets in rats. METHODS: Bone mesenchymal stem cells (BMSCs) were isolated from bone marrow of 3-week SD rats by adherent method. Maxillary posterior teeth of 36 6-week SD rats were extracted and materials were implanted into sockets according to grouping. The rats were divided into 4 groups: compound group with implanting BMSCs-Bio-Oss-bFGF compound, powder group with implanting Bio-Oss, BMSCs group with implanting BMSCs, and control group without implanting any materials. The sockets were scanned by micro-CT 4 weeks, 12 weeks and 24 weeks after implantation. Two-way ANOVA was used to assess whether there was significant difference between groups with GraphPad Prism 6.0 software package. RESULTS: There was no significant difference among groups in bone mineral density (BMD), trabecular separation(Tb.Sp), trabecular thickness(Tb.Th), degree of anisotropy(DA), and trabecular number(Tb.N) 4 weeks after implantation. By 12 weeks, BMD of compound group was significantly greater than those of BMSCs group, powder group and control group (P＜0.05), and significantly greater than those of powder group and control group at 24 weeks (P＜0.05). Tb.Th of compound group was significantly greater than that of BMSCs group at 12 and 24 weeks(P＜0.05). DA had no significant difference among groups at 4, 12, and 24 weeks (P＞0.05). Tb.Sp of compound group was significantly smaller than those of powder group, BMSCs group and control group at 24 weeks(P＜0.05). Tb.N of compound group was significantly greater than those of BMSCs group and control group(P＜0.05). CONCLUSIONS: The compound of rat allogeneic BMSCs-Bio-Oss-bFGF improves socket healing.

Molecular characterization, expression and functional analysis of large yellow croaker (Larimichthys crocea) peroxisome proliferator-activated receptor gamma.

The peroxisome proliferator-activated receptor gamma (PPARγ) are nuclear receptors with distinct roles in energy metabolism and immunity. Although extensively studied in mammals, immunomodulatory roles of this molecule in teleost fish remain to be investigated. In this study, large yellow croaker (Larimichthys crocea) PPARγ (LcPPARγ) sequence was cloned, which encodes a polypeptide of 541 amino acids that include signature domains belonging to the nuclear receptor superfamily. Phylogenetically, LcPPARγ was most closely related to PPARγ derived from European sea bass (Dicentrarchus labrax). Quantitative analysis shown a ubiquitous expression of this molecule, with highest expression level detected in the intestine. The expression of LcPPARγ was decreased in the intestine, muscle, body kidney, spleen and head kidney-derived monocytes/macrophages (MO/MФs) over the course of Vibrio alginolyticus (V. alginolyticus) infection. In contrast, an up-regulation of LcPPARγ was observed in head kidney-derived MO/MФs following docosahexaenoic acid (DHA) treatment. This increase in LcPPARγ leads to an up-regulation of LcCD11b and LcCD18 and an enhancement of complement-mediated phagocytosis. Furthermore, cytokine secretions of V. alginolyticus-stimulated MO/MФs were modulated following LcPPARγ activations that up-regulated the expression of LcIL-10, while decreased the expression of LcIL-1ß, LcTNF-α and LcTGF-ß1. Overall, our results indicated that LcPPARγ plays a role in regulating functions of MO/MФs and likely contribute to MO/MФs polarization.

The effect of new preoperative preparation method compared to conventional method in complex acetabular fractures: minimum 2-year follow-up.

OBJECTIVE: This study aimed to compare the efficacy and safety of the new method including 3D printing-based preoperative planning, surgical workshop, and contouring of the plate versus conventional method in the surgical treatment of complex acetabular fractures. METHODS: We retrospectively analyzed the data in a cohort of 88 patients of complex acetabular fracture with mean 29.95 ± 4.84 months (24-41 months) follow-up. Patients were divided into two groups. Group 1 consisting of 41 patients were performed previewed surgery with a 3D printing-based pre-contoured plate on a 3D printing model. Group 2, comprised of 47 patients, were treated by the traditional contoured plate technique. The quality of reduction was assessed using criteria described by Matta. Functional outcome was evaluated using Modified Postel Merle D'Aubigne score. A custom-made quiz was used to evaluate the chief assistant. RESULTS: The study showed no significant differences in measured preoperative variables except for the age between the Group 1 and Group 2 (p > 0.05). Compared with the Group 2, the intraoperative blood loss, operative time was significantly decreased in Group 1 (p < 0.05). There were no significant statistical differences in the quality of reduction and Modified Postel Merle D'Aubigne score (p > 0.05). The result of evaluation of assistant in Group 1 was significantly high than in Group 2 (p < 0.05). CONCLUSION: 3D printing-based pre-contoured plate is a more effective and reliable method than traditional contoured plate technique for treating the complex acetabular fractures. Meanwhile, the 3D printing is a useful orthopedic surgical education tool which can improve the understanding of the complex acetabular fracture for a young surgeon.

Deep Vein Thrombosis in Patients with Intertrochanteric Fracture: A Retrospective Study.

BACKGROUND: This study aimed to investigate the incidences of pre- and post-operative lower extremity deep venous thrombosis (DVT) in hospitalized patients with intertrochanteric fractures and to analyze the relevant risk factors. METHODS: A retrospective study was conducted between July 2014 and October 2016 in 218 intertrochanteric fracture patients who presented at Xi'an Honghui Hospital and underwent Doppler ultrasonography for DVT diagnosis. We divided DVT into distal, proximal, and mixed thrombosis. Patients were divided into either the thrombosis or no thrombosis group according to preoperative and postoperative ultrasonography results. All patients were evaluated for the risk factors associated with thrombosis. RESULTS: A total of 37.61% of preoperative patients had DVT, and the postoperative incidence increased to 58.72%. The days between fracture and hospitalization and the days between fracture and surgery were independent risk factors for preoperative DVT. The days between fracture and hospitalization and d-dimer levels at postoperative 1 day were independent risk factors of postoperative DVT. In total 23.4% of the patients progressed from having no thrombosis preoperatively to having distal, proximal, or mixed DVT postoperatively (22.02%, 0.46%, and 0.92%, respectively). Distal DVT constituted 86.59% and 90.63% of all preoperative and postoperative DVTs, respectively. CONCLUSION: Intertrochanteric fracture is a common type of hip fracture in the elderly, and the incidence of DVT after intertrochanteric fracture may be underestimated. Early intervention (early admission and early surgery) might reduce the incidence of DVT.

Deep Vein Thrombosis in the Uninjured Limb in Patients with Lower Extremity Fractures: A Retrospective Study.

OBJECTIVE: This study is aimed at investigating the incidence of deep vein thrombosis (DVT) in the uninjured limb during hospitalization and 1 month after surgery in patients with lower extremity fractures. METHODS: We collected the clinical data of patients with lower extremity fractures in Xi'an Honghui Hospital. Doppler ultrasonography was used to diagnose DVT. According to the results of ultrasonography, the patients were divided into two groups: uninjured limb with DVT group and uninjured limb without DVT group. RESULTS: A total of 494 patients who met all inclusion criteria were included in this study. The incidence rate of DVT in the uninjured limb was 19.84% and 18.83% during hospitalization and 1 month after surgery, respectively. Age (OR = 1.035, 95% CI: 1.013-1.059; P = 0.002) and D-dimer level 1 day after surgery (OR = 1.065, 95% CI: 1.030-1.102; P < 0.001) were independent risk factors for DVT during hospitalization. Similarly, age (OR = 1.045, 95% CI: 1.021-1.070; P < 0.001) and D-dimer level 1 day after surgery (OR = 1.048, 95% CI: 1.014-1.083; P = 0.006) were independent risk factors for DVT 1 month after surgery. During hospitalization and 1 month after surgery, 15.79% and 12.35% of patients had double lower limb thrombosis and 4.04% and 6.48% of patients had DVT in the uninjured limb only, respectively. CONCLUSION: The actual incidence of DVT in the uninjured limb in patients with lower extremity fractures cannot be ignored despite the use of anticoagulants for prevention or treatment during hospitalization. We should also be aware of DVT in the uninjured limb while focusing on DVT in the injured limb.

Potential influence of COVID-19/ACE2 on the female reproductive system.

The 2019 novel coronavirus (2019-nCoV) appeared in December 2019 and then spread throughout the world rapidly. The virus invades the target cell by binding to angiotensin-converting enzyme (ACE) 2 and modulates the expression of ACE2 in host cells. ACE2, a pivotal component of the renin-angiotensin system, exerts its physiological functions by modulating the levels of angiotensin II (Ang II) and Ang-(1-7). We reviewed the literature that reported the distribution and function of ACE2 in the female reproductive system, hoping to clarify the potential harm of 2019-nCoV to female fertility. The available evidence suggests that ACE2 is widely expressed in the ovary, uterus, vagina and placenta. Therefore, we believe that apart from droplets and contact transmission, the possibility of mother-to-child and sexual transmission also exists. Ang II, ACE2 and Ang-(1-7) regulate follicle development and ovulation, modulate luteal angiogenesis and degeneration, and also influence the regular changes in endometrial tissue and embryo development. Taking these functions into account, 2019-nCoV may disturb the female reproductive functions through regulating ACE2.

Deep vein thrombosis in the lower extremities after femoral neck fracture: A retrospective observational study.

PURPOSE: The actual incidence of deep vein thrombosis (DVT) in femoral neck fractures is underestimated. This study aimed to investigate the incidence of DVT in the lower extremities after femoral neck fracture before and after operation. METHODS: The clinical data of patients with femoral neck fractures treated at Xi'an Honghui Hospital between July 1, 2016, and December 31, 2018, were collected. The patients were examined with ultrasonography before and after operation and divided into thrombosis and non-thrombosis groups according to their ultrasonographic results. The incidence of DVT was reported as a percentage. RESULTS: The incidence rates of preoperative and postoperative DVT were 32% and 56%, respectively. DVT on the uninjured side constituted 45% of all preoperative DVT and 43% of all postoperative DVT. Peripheral DVT constituted 90% and 84% of all preoperative and postoperative DVT, respectively. Diabetes was an independent risk factor of preoperative DVT. Blood loss was an independent risk factor of postoperative DVT, and open reduction and internal fixation surgical procedure was independent protective factor of postoperative DVT as compared with hemiarthroplasty and total hip replacement. CONCLUSIONS: The incidence rates of preoperative and postoperative DVT in the patients with femoral neck fracture were high, and orthopedists should pay more attention to DVT as a complication.

[Effect of low-intensity pulsed ultrasound combined with triamcinolone acetonide on oral mucosal ulcer healing in syrian hamster].

PURPOSE: To evaluate the effect of low-intensity pulsed ultrasound（LIPUS） combined with triamcinolone acetonide on oral mucosal ulcer in syrian hamster in several ways, including healing time, contents of superoxide dismutase（SOD）and malondialdehyde (MDA). METHODS: Sixty syrian hamsters were randomly divided into 5 groups, including a baseline group (containing a normal baseline group and a model baseline group, n=6) and 4 experimental groups (LIPUS processing and drug use group, LIPUS group, drug group and a normal control group without any processing, n=12). Four experimental groups and model baseline group were given oxygen free radicals to model the oral mucosal ulcer. At 24 h after the last treatment, the healing time of ulcer, content of SOD and MDA were compared between each group. SPSS 20.0 software package was used for statistical analysis. RESULTS: Compared with LIPUS group,drug group and control group, the healing time of oral mucosal ulcer in LIPUS and drug combined group was shortened. At 24 h after the last treatment, the activity of SOD showed that the LIPUS and drug combined group[(2.32±0.30) U/mgprot] were significantly higher than the model baseline group[(1.48±0.29) U/mgprot], the LIPUS group[(1.83±0.15) U/mgprot], the drug group[(1.76±0.25) U/mgprot] and control group[(1.71±0.18) U/mgprot] (P<0.05). The results of MDA content showed that the LIPUS and drug combined group [(8.17±0.21) nmol/mgprot] were significantly lower than the model baseline group[(9.41±0.22) nmol/mgprot], the LIPUS group[(9.00±0.44) nmol/mgprot], the drug group [(9.04±0.43) nmol/mgprot] and control group[(9.03±0.46) nmol/mgprot] (P<0.05). After oral mucosal ulcer healing, the activity of SOD and MDA showed that the LIPUS and drug combined group, the LIPUS group, the drug group and control group were not significantly different from the normal baseline group (P>0.05). CONCLUSIONS: Low-intensity pulsed ultrasound combined with triamcinolone acetonide can effectively improve the activity of SOD and reduce the contents of MDA in ulcerated tissues, and therefore accelerate the process of ulcer healing．.

BMP4 inhibits myogenic differentiation of bone marrow-derived mesenchymal stromal cells in mdx mice.

BACKGROUND AIMS: Bone marrow-derived mesenchymal stromal cells (BMSCs) are a promising therapeutic option for treating Duchenne muscular dystrophy (DMD). Myogenic differentiation occurs in the skeletal muscle of the mdx mouse (a mouse model of DMD) after BMSC transplantation. The transcription factor bone morphogenic protein 4 (BMP4) plays a crucial role in growth regulation, differentiation and survival of many cell types, including BMSCs. We treated BMSCs with BMP4 or the BMP antagonist noggin to examine the effects of BMP signaling on the myogenic potential of BMSCs in mdx mice. METHODS: We added BMP4 or noggin to cultured BMSCs under myogenic differentiation conditions. We then injected BMP4- or noggin-treated BMSCs into the muscles of mdx mice to determine their myogenic potential. RESULTS: We found that the expression levels of desmin and myosin heavy chain decreased after treating BMSCs with BMP4, whereas the expression levels of phosphorylated Smad, a downstream target of BMP4, were higher in these BMSCs than in the controls. Mdx mouse muscles injected with BMSCs pretreated with BMP4 showed decreased dystrophin expression and increased phosphorylated Smad levels compared with muscles injected with non-treated BMSCs. The opposite effects were seen after pretreatment with noggin, as expected. CONCLUSIONS: Our results identified BMP/Smad signaling as an essential negative regulator of promyogenic BMSC activity; inhibition of this pathway improved the efficiency of BMSC myogenic differentiation, which suggests that this pathway might serve as a target to regulate BMSC function for better myogenic differentiation during treatment of DMD and degenerative skeletal muscle diseases.

MicroRNA-370 Attenuates Hepatic Fibrogenesis by Targeting Smoothened.

BACKGROUND AND AIMS: Recent research shows that abnormal expression of microRNA plays an important role in the process of hepatic fibrosis . miR-370 has been reported to be involved in liver function and is suppressed during hepatic carcinogenesis. The aim of this study was to investigate the role of miR-370 in hepatic fibrosis. METHODS: The expression levels of miR-370 in rat fibrotic livers and activated hepatic stellate cells (HSCs) were evaluated by quantitative real-time PCR. The effect of miR-370 on the activation of HSCs was analyzed by flow cytometric analyses, real-time PCR and Western blot. Adenovirus carrying miR-370 was injected through the tail vein to access the effect of miR-370 on hepatic fibrosis induced by CCl4 in rats. The downstream targets of miR-370 were predicted by the Target Scan database and verified by luciferase assays, real-time PCR and Western blot in HSCs and were further confirmed by immunohistochemistry in vivo. RESULTS: Real-time PCR showed that miR-370 expression was significantly reduced in rat fibrotic livers and TGFß1-stimulated HSCs. Overexpression of miR-370 inhibited the proliferation of HSC-T6 cells via inducing cell apoptosis and suppressed the activation of HSCs. Upregulation of miR-370 obviously attenuated the CCl4-induced liver fibrosis in rats. miR-370 was directly bound to the 3'UTR of Smoothened (SMO) and suppressed the expression of SMO in HSCs and fibrotic livers. CONCLUSIONS: Our study demonstrated that miR-370 plays an inhibitory role in hepatic fibrogenesis by targeting SMO. Restoration of miR-370 may have beneficial effects on the treatment of liver fibrosis.

Detection of the human endogenous retrovirus ERV3-encoded Env-protein in human tissues using antibody-based proteomics.

OBJECTIVES: There is growing evidence to suggest that human endogenous retroviruses (HERVs) have contributed to human evolution, being expressed in development, normal physiology and disease. A key difficulty in the scientific evaluation of this potential viral contribution is the accurate demonstration of virally expressed protein in specific human cells and tissues. In this study, we have adopted the endogenous retrovirus, ERV3, as our test model in developing a reliable high-capacity methodology for the expression of such endogenous retrovirus-coded protein. DESIGN: Two affinity-purified polyclonal antibodies to ERV3 Env-encoded protein were generated to detect the corresponding protein expression pattern in specific human cells, tissues and organs. PARTICIPANTS: Sampling included normal tissues from 144 individuals ranging from childhood to old age. This included more than forty different tissues and organs and some 216 different cancer tissues representing the twenty commonest forms of human cancer. SETTING: The Rudbeck Laboratory, Uppsala University and Uppsala University Hospital, Uppsala, Sweden. MAIN OUTCOME MEASURES: The potential expression at likely physiological level of the ERV3Env encoded protein in a wide range of human cells, tissues and organs. RESULTS: We found that ERV3 encoded Env protein is expressed at substantive levels in placenta, testis, adrenal gland, corpus luteum, Fallopian tubes, sebaceous glands, astrocytes, bronchial epithelium and the ducts of the salivary glands. Substantive expression was also seen in a variety of epithelial cells as well as cells known to undergo fusion in inflammation and in normal physiology, including fused macrophages, myocardium and striated muscle. This contrasted strongly with the low levels expressed in other tissues types. These findings suggest that this virus plays a significant role in human physiology and may also play a possible role in disease. CONCLUSION: This technique can now be extended to the study of other HERV genomes within the human chromosomes that may have contributed to human evolution, physiology and disease.