RNA-binding proteins potentially regulate the alternative splicing of apoptotic genes during knee osteoarthritis progression.

BACKGROUND: Alternative splicing (AS) is a principal mode of genetic regulation and one of the most widely used mechanisms to generate structurally and functionally distinct mRNA and protein variants. Dysregulation of AS may result in aberrant transcription and protein products, leading to the emergence of human diseases. Although considered important for regulating gene expression, genome-wide AS dysregulation, underlying mechanisms, and clinical relevance in knee osteoarthritis (OA) remain unelucidated. Therefore, in this study, we elucidated and validated AS events and their regulatory mechanisms during OA progression. RESULTS: In this study, we identified differentially expressed genes between human OA and healthy meniscus samples. Among them, the OA-associated genes were primarily enriched in biological pathways such as extracellular matrix organization and ossification. The predominant OA-associated regulated AS (RAS) events were found to be involved in apoptosis during OA development. The expression of the apoptosis-related gene BCL2L13, XAF1, and NF2 were significantly different between OA and healthy meniscus samples. The construction of a covariation network of RNA-binding proteins (RBPs) and RAS genes revealed that differentially expressed RBP genes LAMA2 and CUL4B may regulate the apoptotic genes XAF1 and BCL2L13 to undergo AS events during OA progression. Finally, RT-qPCR revealed that CUL4B expression was significantly higher in OA meniscus samples than in normal controls and that the AS ratio of XAF1 was significantly different between control and OA samples; these findings were consistent with their expected expression and regulatory relationships. CONCLUSIONS: Differentially expressed RBPs may regulate the AS of apoptotic genes during knee OA progression. XAF1 and its regulator, CUL4B, may serve as novel biomarkers and potential therapeutic targets for this disease.

Single-cell transcriptome analyses reveal critical regulators of spermatogonial stem cell fate transitions.

BACKGROUND: Spermatogonial stem cells (SSCs) are the foundation cells for continual spermatogenesis and germline regeneration in mammals. SSC activities reside in the undifferentiated spermatogonial population, and currently, the molecular identities of SSCs and their committed progenitors remain unclear. RESULTS: We performed single-cell transcriptome analysis on isolated undifferentiated spermatogonia from mice to decipher the molecular signatures of SSC fate transitions. Through comprehensive analysis, we delineated the developmental trajectory and identified candidate transcription factors (TFs) involved in the fate transitions of SSCs and their progenitors in distinct states. Specifically, we characterized the Asingle spermatogonial subtype marked by the expression of Eomes. Eomes+ cells contained enriched transplantable SSCs, and more than 90% of the cells remained in the quiescent state. Conditional deletion of Eomes in the germline did not impact steady-state spermatogenesis but enhanced SSC regeneration. Forced expression of Eomes in spermatogenic cells disrupted spermatogenesis mainly by affecting the cell cycle progression of undifferentiated spermatogonia. After injury, Eomes+ cells re-enter the cell cycle and divide to expand the SSC pool. Eomes+ cells consisted of 7 different subsets of cells at single-cell resolution, and genes enriched in glycolysis/gluconeogenesis and the PI3/Akt signaling pathway participated in the SSC regeneration process. CONCLUSIONS: In this study, we explored the molecular characteristics and critical regulators of subpopulations of undifferentiated spermatogonia. The findings of the present study described a quiescent SSC subpopulation, Eomes+ spermatogonia, and provided a dynamic transcriptional map of SSC fate determination.

Pancreatic lipase-related protein 2 is selectively expressed by peritubular myoid cells in the murine testis and sustains long-term spermatogenesis.

Spermatogenesis is a complicated process of germ cell differentiation that occurs within the seminiferous tubule in the testis. Peritubular myoid cells (PTMCs) produce major components of the basement membrane that separates and ensures the structural integrity of seminiferous tubules. These cells secrete niche factors to promote spermatogonial stem cell (SSC) maintenance and mediate androgen signals to direct spermatid development. However, the regulatory mechanisms underlying the identity and function of PTMCs have not been fully elucidated. In the present study, we showed that the expression of pancreatic lipase-related protein 2 (Pnliprp2) was restricted in PTMCs in the testis and that its genetic ablation caused age-dependent defects in spermatogenesis. The fertility of Pnliprp2 knockout animals (Pnliprp2-/-) was normal at a young age but declined sharply beginning at 9 months. Pnliprp2 deletion impaired the homeostasis of undifferentiated spermatogonia and severely disrupted the development and function of spermatids. Integrated analyses of single-cell RNA-seq and metabolomics data revealed that glyceride metabolism was changed in PTMCs from Pnliprp2-/- mice. Further analysis found that 60 metabolites were altered in the sperm of the Pnliprp2-/- animals; notably, lipid metabolism was significantly dysregulated. Collectively, these results revealed that Pnliprp2 was exclusively expressed in PTMCs in the testis and played a novel role in supporting continual spermatogenesis in mice. The outcomes of these findings highlight the function of lipid metabolism in reproduction and provide new insights into the regulation of PTMCs in mammals.

Receptive Field Space for Point Cloud Analysis.

Similar to convolutional neural networks for image processing, existing analysis methods for 3D point clouds often require the designation of a local neighborhood to describe the local features of the point cloud. This local neighborhood is typically manually specified, which makes it impossible for the network to dynamically adjust the receptive field's range. If the range is too large, it tends to overlook local details, and if it is too small, it cannot establish global dependencies. To address this issue, we introduce in this paper a new concept: receptive field space (RFS). With a minor computational cost, we extract features from multiple consecutive receptive field ranges to form this new receptive field space. On this basis, we further propose a receptive field space attention mechanism, enabling the network to adaptively select the most effective receptive field range from RFS, thus equipping the network with the ability to adjust granularity adaptively. Our approach achieved state-of-the-art performance in both point cloud classification, with an overall accuracy (OA) of 94.2%, and part segmentation, achieving an mIoU of 86.0%, demonstrating the effectiveness of our method.

Clinical application of modified Crain classification in the Design of Anterior Cruciate Ligament Reconstruction with remnant preservation.

BACKGROUND: To investigate the clinical application of modified Crain classification in anterior cruciate ligament (ACL) reconstruction (ACLR) with remnant preservation. METHODS: The subjects were 70 patients with ACL injury who underwent ACLR from May 2016 to June 2018, and their general data were recorded. They were randomly divided into modified remnant-preserved ACLR group (group M, n = 35) and non remnant-preserved ACLR group (group N, n = 35). ACLR program with remnant preservation was designed based on modified Crain classification in group M, while ACL remnants were completely cleaned during ACLR in group N. Subsequently, the two groups were compared in terms of operation time, complications, as well as Lysholm score, international knee documentation committee (IKDC) score and positive rate of Lachman test of knee joint before operation and at 3, 6 and 12 months after operation. RESULTS: Both the groups showed good postoperative efficacy, and none had complications like limited knee extension or cyclops lesion. The comparison results found that group M (72.49 ± 7.64 min) required longer operation time than group N (66.06 ± 6.37 min) (P < 0.05). Lysholm score and IKDC score at 3, 6 and 12 months after operation in the two groups were significantly higher than those before operation (P < 0.05); group M had higher Lysholm score and IKDC score at 3 months and 6 months after operation compared with group N (P < 0.05). Additionally, the positive rate of Lachman test at 3, 6 and 12 months after operation in both groups was significantly lower than that before operation (P < 0.05), but there was no significant difference between group M and group N. CONCLUSION: With the modified Crain classification, many remnant-preserved reconstruction techniques can be rationally used to completely preserve the remnant ligament tissue during operation and improve knee joint function and joint stability with few complications.

Paternal hypoxia exposure impairs fertilization process and preimplantation embryo development.

Environmental hypoxia exposure causes fertility problems in human and animals. Compelling evidence suggests that chronic hypoxia impairs spermatogenesis and reduces sperm motility. However, it is unclear whether paternal hypoxic exposure affects fertilization and early embryo development. In the present study, we exposed male mice to high altitude (3200 m above sea level) for 7 or 60 days to evaluate the effects of hypoxia on sperm quality, zygotic DNA methylation and blastocyst formation. Compared with age-matched controls, hypoxia-treated males exhibited reduced fertility after mating with normoxic females as a result of defects in sperm motility and function. Results of in vitro fertilization (IVF) experiments revealed that 60 days' exposure significantly reduced cleavage and blastocyst rates by 30% and 70%, respectively. Immunohistochemical staining of pronuclear formation indicated that the pronuclear formation process was disturbed and expression of imprinted genes was reduced in early embryos after paternal hypoxia. Overall, the findings of this study suggested that exposing male mice to hypoxia impaired sperm function and affected key events during early embryo development in mammals.

ß-Substituted Alkenyl Heteroarenes as Dipolarophiles in the Cu(I)-Catalyzed Asymmetric 1,3-Dipolar Cycloaddition of Azomethine Ylides Empowered by a Dual Activation Strategy: Stereoselectivity and Mechanistic Insight.

The catalytic asymmetric 1,3-dipolar cycloaddition reactions of azomethine ylides with various electron-deficient alkenes provide the most straightforward protocol for the preparation of enantioenriched pyrrolidines in organic synthesis. However, the employment of conjugated alkenyl heteroarenes as dipolarophiles in such protocols to afford a class of particularly important molecules in medicinal chemistry is still a great challenge. Herein, we report that various ß-substituted alkenyl heteroarenes, challenging internal alkene substrates without a strong electron-withdrawing substituent, were successfully employed as dipolarophiles for the first time in the Cu(I)-catalyzed asymmetric 1,3-dipolar cycloaddition of azomethine ylides. This reaction furnishes a large array of multistereogenic heterocycles incorporating both the biologically important pyrrolidine and heteroarene skeletons in good yields with exclusive diastereoselectivity and excellent enantioselectivity. Our extensive density functional theory (DFT) calculations proposed a working model to explain the origin of the stereochemical outcome and elucidated uncommon dual activation/coordination of both the dipole and dipolarophile substrates by the metal, in which a sterically bulky, rigid, and monodentate phosphoramidite ligand with triple-homoaxial chirality plays a pivotal role in providing an effective chiral pocket around the metal center, resulting in high enantioselectivity. The additional coordination of the heteroatom in the dipolarophile substrate to Cu is also critical for the exclusive diastereoselectivity and enhanced reactivity. Our calculations also predicted the reverse and high enantioinduction for the corresponding substrates with monocyclic heteroarenes as well as regiospecific cycloaddition to the less reactive internal C═C bond of one related dipolarophile diene substrate. Such unique steric effect-directed enantioswitching and coordination-directed regioselectivity were verified experimentally.

Application of Real-Time Augmented Reality Laparoscopic Navigation in Splenectomy for Massive Splenomegaly.

OBJECTIVES: To evaluate the clinical impact and technical feasibility of augmented reality laparoscopic navigation (ARLN) system in laparoscopic splenectomy for massive splenomegaly. METHODS: The clinical data of 17 consecutive patients who underwent laparoscopic splenectomy using ARLN (ARLN group) and 26 patients without ARLN guidance (Non-ARLN group) between January 2018 and April 2020 were enrolled. Propensity score matching (PSM) analysis was performed between the patients with and without ARLN guidance at a ratio of 1:1. RESULTS: Mean intraoperative blood loss was significantly lower in the ARLN-group than in the Non-ARLN group (306.6 ml vs. 462.6 ml, p = 0.047). All the patients in the ARLN-group achieved successful splenic artery dissection, while surgical success was achieved in 12 patients in the Non-ARLN group (p = 0.044). Postoperative hospital stay was significantly longer in the Non-ARLN group (3.8 days vs. 4.5 days, p = 0.040). CONCLUSIONS: ARLN can provide feasible and accurate intraoperative image guidance, and it could be helpful in the performance of laparoscopic splenectomy for massive splenomegaly.

Proscillaridin A induces apoptosis and inhibits the metastasis of osteosarcoma in vitro and in vivo.

The side effects of chemotherapy, drug resistance, and tumor metastasis hinder the development of treatment for osteosarcoma, leading to poor prognosis of patients with the disease. Proscillaridin A, a kind of cardiac glycoside, has been proven to have anti-proliferative properties in many malignant tumors, but the efficacy of the drug in treating osteosarcoma is unclear. In the present study, we assessed the effects of Proscillaridin A on osteosarcoma and investigated its underlying action mechanism. The cell cytotoxicity assay showed that Proscillaridin A significantly inhibited the proliferation of 143B cells in a dose- and time-dependent manner. Also, flow cytometry and invasion assay revealed that Proscillaridin A induced apoptosis and reduced 143B cell motility. Western blotting and PCR were used to detect the expressions of Bcl-xl and MMP2 and showed that mRNA/protein expression levels decreased significantly in Proscillaridin A-treated osteosarcoma cells. Using a mouse xenograft model, we found that Proscillaridin A treatment significantly inhibited tumor growth and lung metastasis in vivo and decreased the expression levels of Bcl-xl and MMP2. No noticeable side effect was observed in the liver, kidney, and hematological functions. Conclusively, Proscillaridin A suppressed proliferation, induced apoptosis, and inhibited 143B cell metastasis in vitro and in vivo, and these effects could be mediated by downregulating the expressions of Bcl-xl and MMP2.

A novel method of fluorescent imaging can guide hepatectomy for intrahepatic cholangiocarcinoma with intrahepatic biliary obstruction.

OBJECTIVE: The aim of this study was to present a novel bile-duct obstructed area imaging (BOAI) and to investigate the feasibility and accuracy of this method in guiding hepatectomy for intrahepatic cholangiocarcinoma (ICC) with intrahepatic biliary obstruction. METHODS: From May 2017 to October 2019, eligible patients who underwent hepatectomy guided by BOAI were enrolled. Perioperative outcomes and operative data were analyzed. To assess the feasibility of BOAI and Glissonean pedicle approach, demarcations based on them were compared. To verify the accuracy of BOAI staining of the target territory, simple linear regression analysis, and intraclass correlation coefficient were used to examine the relationship between predicted resected liver volume (PRLV) and actual resected liver volume (ARLV). RESULTS: BOAI staining achieved valid demarcation in 15 (93.8%) of 16 patients, whereas the ischemic line achieved valid demarcation in only nine patients (57.3%; p = .017). ARLV and PRLV had a strong positive correlation (PRLV = 60.06 + 0.925 × ARLV; R = .945; p = .000). Meanwhile, ARLV (intraclass correlation coefficient = .971) achieved an excellent agreement with PRLV (p < .001). CONCLUSIONS: The novel BOAI staining method can provide valid, feasible, and accurate demarcation line and may be an effective method in the surgical treatment of intrahepatic biliary obstruction.

Comparison of subtotal vertebral resection with reconstruction and percutaneous vertebroplasty for treatment of metastasis in the lumbar spine.

Purpose: Tumor metastasis in the spine can cause pain and fractures, leading to deformities, and deficits in movement, sensation, and bowel/bladder function. Percutaneous vertebroplasty (PVP) and subtotal vertebral resection with reconstruction (SVR) are suitable treatments, but their relative clinical efficacy is uncertain. The purpose of this retrospective cohort study was to compare the management and clinical effect of SVR for lumbar metastatic tumor with PVP.Methods: Sixty-seven patients (mean age: 58.6 years) with metastases in the lumbar spine received SVR or PVP at our institution between 2010 and 2013. Thirty-three patients received SVR via a posterior approach, in which vertebrae were resected, with the anterior and lateral walls retained using polymethylmethacrylate (PMMA), followed by reconstruction and pedicle screw fixation. Thirty-four patients received PVP via the vertebral pedicle. Patients were followed for 3-26 months.Results: None of the patients experienced serious complications after surgery, and all patients experienced significant amelioration of pain. Twelve patients (8 in the PVP group and 4 in the SVR group) died during the follow-up, and the survival time was significantly longer in the SVR group. Two patients in the SVR group and 7 patients in the PVP groups experienced recurrence during follow-up, but the groups had no significant difference in local recurrence. Both treatments significantly reduced scores for pain on a visual analog scale (pain-VAS) and disability (Oswestry Disability Index [ODI]), and increased performance status (Karnofsky Performance Status [KPS]). Compared with the PVP group, the SVR group had better ODI score at 1 month and 3 months after surgery and a higher KPS score at 1 month after surgery. The two groups had no significant difference in pain-VAS scores during follow-up.Conclusions: SVR is a reliable treatment for lumbar metastatic tumor and provides good survival rate and satisfying follow-up results.

The anticancer effects of 2-methoxyestradiol on human huh7 cells in vitro and in vivo.

Hepatocellular carcinoma (HCC) is associated with a poor prognosis. 2-methoxyestradiol (2-ME) is currently under preclinical evaluation as a treatment for many malignancies, but the utility of the drug in terms of HCC treatment remains unclear. Here, we explored the effect of 2-ME on human huh7 cell proliferation and apoptosis and discuss the possible molecular mechanisms involved. The MTT assay showed that proliferation was markedly inhibited by 2-ME (at 5, 10, 15, and 20 µmol/L) in a time- and dose-dependent manner. Moreover, flow cytometry indicated that 2-ME induced cell cycle arrest at the G2/M phase, and early apoptosis. We used Western blotting and PCR to detect the expression of vascular endothelial growth factor (VEGF) and Bcl-2; 2-ME decreased the mRNA/protein expression levels of both effectors. Furthermore, 2-ME remarkably suppressed xenograft tumor growth in nude mice, and no visible toxicity was observed in either the liver or kidneys. Immunohistochemically, the Bcl-2 and VEGF expression levels were significantly lower than those of controls. Thus, 2-ME inhibited huh7 cell proliferation, promoted apoptosis, and suppressed xenograft tumor growth in nude mice, perhaps reflecting the effects of the drug on VEGF and Bcl-2 expression.

Synergistic Cu/Pd-Catalyzed Asymmetric Allenylic Alkylation of Azomethine Ylides for the Construction of α-Allene-Substituted Nonproteinogenic α-Amino Acids.

An unprecedented asymmetric allenylic alkylation of readily available imine esters, which was enabled by a synergistic Cu/Pd catalysis, has been developed. This dual catalytic system possesses good substrate compatibility, delivering a diverse array of nonproteinogenic α-allenylic α-mono- or α,α-disubstituted α-amino acids (α-AAs) with high yields and generally excellent enantioselectivities. Furthermore, the scalability and practicability of the current synthetic protocol were proven by performing gram-scale reactions and by the first catalytic asymmetric synthesis of naturally occurring (S)-γ-allenic α-amino acid, respectively.

Synergistic effect of docetaxel combined with cisplatin on inhibiting human osteosarcoma in nude mice.

Cisplatin (CDDP) has been shown to be a promising anticancer drug that is effective against many types of cancer, which include osteosarcoma (OS). However, its therapeutic application is restricted by its toxicity in normal tissues, side effects caused in patients, and chemotherapy resistance. Thus, to further improve patients' treatment, the development of novel, more effective and well tolerated therapeutic approaches against OS in clinical is urgent and important. In the present study, nude mice were inoculated subcutaneously with injections of HOS8603 cells, CDDP and docetaxel (DTX) were administered intraperitoneally respectively. The inhibitive effects and the side effects were observed. Tumor weights and volumes were significantly lower and the tumor inhibition rate was significantly higher in the combination group than those of either drug alone or vehicle. The cell density in the tumor tissue was significantly decreased, apoptotic and necrotic cell death was significantly increased in the combination group, as compared with those of either drug alone or vehicle. In addition, there was no obvious side effect happening besides the appearance of erythema and papules in some mice. These results suggest that the combined effects of CDDP and DTX on the growth of human OS in vivo were superior to the single effects. CDDP combined with DTX had synergistic effects at lower concentrations and promoted apoptosis, but did not increase the side effects of chemotherapy.

Enhanced Li Storage Stability Induced by Locating Sn in Metal-Organic Frameworks.

By locating elemental Sn in an open anionic framework, the particle cracking arising from huge volume expansion of Sn-based anode materials during lithiation/delithiation is alleviated, and the cycling stability is greatly improved. The Sn-based metal-organic framework anode material shows superior cyclic stability, with a capacity retention >92 % (after 200 cycles) and high lithium storage capacity (610 mAh g-1 ).

Accuracy of actual resected liver volume in anatomical liver resections guided by 3-dimensional parenchymal staining using fusion indocyanine green fluorescence imaging.

BACKGROUND: The aim of this study was to assess the accuracy of actual resected liver volume (ARLV) in anatomical liver resections (ALRs) guided by 3-dimensional parenchymal staining using fusion indocyanine green fluorescence imaging (IGFI). METHODS: Patients eligible for hepatic resection were enrolled in the current study from January 2016 to November 2017. All patients underwent surgery planning based on Medical Image Three-Dimensional Visualization System (MI-3DVS) before the operation, in which predicted resected liver volumes (PRLVs) were calculated. Under 3-dimensional guidance by fusion IGFI, ALRs were performed and ARLVs were measured. Simple linear regression, intra-class correlation coefficient (ICC) and Bland-Altman analysis were used to evaluate the relationship and agreement between PRLV and ARLV. RESULTS: Of the 27 patients who achieved valid demarcation by fusion IGFI, 12 (44.4%) received hemihepatectomy, while 10 (37.0%) and five (18.5%) underwent sectionectomy and segmentectomy, respectively. The relationship and agreement between PRLV (481.37 ± 189.47 cm³) and ARLV (450.57 ± 205.19 cm³) were then evaluated. The simple regression equation obtained was PRLV = 0.874 × ARLV + 87.75 (R = 0.946; P = 0.000). Meanwhile, ARLV (ICC = 0.943) achieved an excellent agreement with PRLV ( P < 0.001); 25 of 27 dots were in the range of 95% confidence interval in Bland-Altman analysis. CONCLUSIONS: In the study, these findings validated the consistency between PRLV calculated by MI-3DVS and ARLV guided by fusion IGFI, which proved that IGFI can accurately guide anatomical hepatectomy. Generally, fusion IGFI can provide a valid, feasible and accurate demarcation line, which can confer precision to hepatic resection.

Eosin†Y as a Direct Hydrogen-Atom Transfer Photocatalyst for the Functionalization of C-H Bonds.

Eosin Y, a well-known economical alternative to metal catalysts in visible-light-driven single-electron transfer-based organic transformations, can behave as an effective direct hydrogen-atom transfer catalyst for C-H activation. Using the alkylation of C-H bonds with electron-deficient alkenes as a model study revealed an extremely broad substrate scope, enabling easy access to a variety of important synthons. This eosin Y-based photocatalytic hydrogen-atom transfer strategy is promising for diverse functionalization of a wide range of native C-H bonds in a green and sustainable manner.

Copper(I)-Catalyzed Asymmetric Desymmetrization through Inverse-Electron-Demand aza-Diels-Alder Reaction: Efficient Access to Tetrahydropyridazines Bearing a Unique α-Chiral Silane Moiety.

An unprecedented copper(I)-catalyzed asymmetric desymmetrization of 5-silylcyclopentadienes with in situ formed azoalkene was realized through an inverse-electron-demand aza-Diels-Alder reaction (IEDDA) pathway, in which 5-silylcyclopentadienes served as efficient enophiles. This new protocol provides a facile access to the biologically important heterocyclic tetrahydropyridazines containing a unique α-chiral silane motif and three adjoining stereogenic centers in generally good yield (up to 92 %) with exclusive regioselectivity, high diastereoselectivity (>20:1 diastereomeric ratio), and excellent enantioselectivity (up to 98 % enantiomeric excess). DFT calculations and control experiments further confirmed the proposed reaction mechanism.

Silver(I)-Catalyzed Atroposelective Desymmetrization of N-Arylmaleimide via 1,3-Dipolar Cycloaddition of Azomethine Ylides: Access to Octahydropyrrolo[3,4-c]pyrrole Derivatives.

A highly efficient Ag(I)-catalyzed atroposelective desymmetrization of N-(2-t-butylphenyl)maleimide via 1,3-dipolar cycloaddition of in situ generated azomethine ylides has been established successfully, affording a facile access to a series of biologically important and enantioenriched octahydropyrrolo[3,4-c]pyrrole derivatives in generally high yields (up to 99%) with excellent levels of diastereo-/enantioselectivities (up to 99% ee, >20:1 dr). Subsequent transformations led to fascinating 2H-pyrrole and polysubstituted pyrrole compounds without loss of stereoselectivity. The absolute configuration of the generated chiral axis has been unambiguously identified as (M) through single-crystal X-ray diffraction analysis. Furthermore, on the basis of the comprehensive experimental results and the absolute configuration of one of the cycloadducts, the origin of the stereoselectivity was proposed to be attributed to the steric congestion imposed by the bulky PPh2 group of the chiral ligand and the tert-butyl group of N-(2-t-butylphenyl)maleimide. The possible hydrogen bond interaction between the NH2 group of the chiral ligand and one of the carbonyl groups of N-(2-t-butylphenyl)maleimide is considered to facilitate stabilizing the transition state.