Sympathetic hyperinnervation drives abdominal aortic aneurysm development by promoting vascular smooth muscle cell phenotypic switching.

INTRODUCTION: Sympathetic hyperinnervation plays an important role in modulating the vascular smooth muscle cell (VSMC) phenotype and vascular diseases, but its role in abdominal aortic aneurysm (AAA) is still unknown. OBJECTIVES: This study aimed to investigate the role of sympathetic hyperinnervation in promoting AAA development and the underlying mechanism involved. METHODS: Western blotting and immunochemical staining were used to detect sympathetic hyperinnervation. We performed sympathetic denervation through coeliac ganglionectomy (CGX) and 6-OHDA administration to understand the role of sympathetic hyperinnervation in AAA and investigated the underlying mechanisms through transcriptome and functional studies. Sema4D knockout (Sema4D-/-) mice were utilized to determine the involvement of Sema4D in inducing sympathetic hyperinnervation and AAA development. RESULTS: We observed sympathetic hyperinnervation, the most important form of sympathetic neural remodeling, in both mouse AAA models and AAA patients. Elimination of sympathetic hyperinnervation by CGX or 6-OHDA significantly inhibited AAA development and progression. We further revealed that sympathetic hyperinnervation promoted VSMC phenotypic switching in AAA by releasing extracellular ATP (eATP) and activating eATP-P2rx4-p38 signaling. Moreover, single-cell RNA sequencing revealed that Sema4D secreted by osteoclast-like cells induces sympathetic nerve diffusion and hyperinnervation through binding to Plxnb1. We consistently observed that AAA progression was significantly ameliorated in Sema4D-deficient mice. CONCLUSIONS: Sympathetic hyperinnervation driven by osteoclast-like cell-derived Sema4D promotes VSMC phenotypic switching and accelerates pathological aneurysm progression by activating the eATP/P2rx4/p38 pathway. Inhibition of sympathetic hyperinnervation emerges as a potential novel therapeutic strategy for preventing and treating AAA.

Asiaticoside alleviates lipopolysaccharide-induced acute lung injury by blocking Sema4D/CD72 and inhibiting mitochondrial dysfunction in RAW264.7 cell and mice.

Acute lung injury (ALI) is a common disease with complex pathogenesis. However, the treatment is mainly symptomatic with limited clinical options. Asiaticoside (AS), a Chinese herbal extract, has protective effects against LPS-induced ALI in mice and inhibits nitric oxide and prostaglandin E2 synthesis; however, the specific mechanism of AS in the prevention and treatment of LPS-induced ALI needs further study. Sema4D/CD72 pathway, mitochondrial dysfunction, and miRNA-21 are closely associated with inflammation. Therefore, the present study aimed to explore whether AS exerts its therapeutic effect on ALI by influencing Sema4D/CD72 pathway and mitochondrial dysfunction, restoring the balance of inflammatory factors, and influencing miRNA-21 expression. Cell and animal experiments were performed to investigate the effect of AS on ALI. Lipopolysaccharide (LPS) was used to establish the ALI model. CCK8 and flow cytometry were used to detect the cell viability and apoptosis rate. HE staining and wet-to-dry weight ratio (W/D) of lung tissue were determined. The expressions of Sema4D, CD72, NF-κB p65, Bax, Bcl2, and caspase 3 in RAW264.7 cells and lung tissues were detected by western blot, and the levels of IL-10 and IL-1ß induced by LPS in supernatant of RAW264.7 cells and BALF were measured by ELISA. And the expression of miRNA-21 in cells and lung tissues was detected by fluorescence quantitative PCR. The result shows that AS treatment suppressed LPS-induced cell damage and lung injury in mice. AS treatment could alleviate the pathological changes such as inflammatory infiltration and histopathological changes in the lungs caused by LPS, and reduce the ratio of W/D. AS significantly alleviated the decrease of mitochondrial membrane potential induced by LPS, inhibited the increase of ROS production, and reduced the expression of mitochondrial fission proteins Drp1 and Fis1. The high-dose AS group significantly downregulated the expression of Sema4D, CD72, phosphorylated NF-κB p65, and apoptosis-related proteins, decreased the pro-inflammatory factor IL-1ß, and enhanced the level of anti-inflammatory factor IL-10. In addition, AS promoted miRNA-21 expression. These effects inhibited apoptosis and restored the balance between anti- and pro-inflammatory factors. This represents the inaugural report elucidating the mechanism by which AS inhibits the Sema4D/CD72 signaling pathway. These findings offer novel insights into the potential application of AS in both preventing and treating ALI.

Genome-wide Analysis Identified SEMA4D, Novel Candidate Gene for Temperature Sensitivity in Patients With Non-Small Cell Lung Cancer.

BACKGROUND: In the era of precision medicine, individual temperature sensitivity has been highlighted. This trait has traditionally been used for cold-heat pattern identification to understand the inherent physical characteristics, which are influenced by genetic factors, of an individual. However, genome-wide association studies (GWASs) on this trait are limited. METHODS: Using genotype data from 90 patients with advanced non-small cell lung cancer (NSCLC) and epidermal growth factor receptor mutations, we performed a GWAS to assess the association between single nucleotide polymorphisms (SNPs) and temperature sensitivity, such as cold and heat scores. The score of each participant was evaluated using self-administered questionnaires on common symptoms and a 15-item symptom-based cold-heat pattern identification questionnaire. RESULTS: The GWAS was adjusted for confounding factors, including age and sex, and significant associations were identified for cold and heat scores: SNP rs145814326, located on the intron of SORCS2 at chromosome 4p16.1, had a P-value of 1.86 × 10-7; and SNP rs79297667, located upstream from SEMA4D at chromosome 9q22.2, had a P-value of 8.97 × 10-8. We also found that the genetic variant regulates the expression level of SEMA4D in the main tissues, including the lungs and white blood cells, in NSCLC. CONCLUSIONS: SEMA4D was found to be significantly associated with temperature sensitivity in patients with NSCLC, suggesting an increased expression of SEMA4D in patients with higher heat scores. The potential role of temperature sensitivity as a prognostic or predictive marker of immune response in NSCLC should be further studied.

D-alanine suppressed osteoclastogenesis derived from bone marrow macrophages and downregulated ERK/p38 signalling pathways.

OBJECTIVES: D-alanine is a residue of the backbone structure of Type Ⅰ Lipoteichoic acid (LTA), which is a virulence factor in inflammation caused by gram-positive bacteria. However, the role of D-alanine in infectious bone destruction has not been investigated. We aimed to explore the role of D-alanine in the proliferation, apoptosis, and differentiation of osteoclasts. DESIGN: Mouse bone marrow-derived macrophages (BMMs) were isolated as osteoclast precursors and stimulated with D-alanine. Cell proliferation and apoptosis were detected using CCK-8 and flow cytometry, respectively. The formation of osteoclasts morphologically observed by tartrate-resistant acid phosphatase staining (TRAP) and immunofluorescence staining. The expressions of osteoclastogenic genes were measured by real-time RT-PCR. The protein expressions of osteoclastogenic markers, p38, and ERK1/2 MAPK signalling were measured by western blot. The expression level of soluble Sema4D was detected via enzyme-linked immunosorbent assay (ELISA). RESULTS: The cell proliferation of BMMs was significantly inhibited by D-alanine in a dose-dependent manner. Apoptosis of BMMs was markedly activated with the stimulation of D-alanine. The differentiation of BMMs into osteoclasts was significantly inhibited by D-alanine, and the gene and protein expressions of NFATc1, c-Fos, and Blimp decreased. Western blot showed that D-alanine inhibited the phosphorylated p38 and ERK1/2 signalling pathways of BMMs. Moreover, the expression level of soluble Sema4D significantly decreased in the supernatant of BMMs due to the D-alanine intervention. CONCLUSION: D-alanine plays a pivotal role in the inhibition of RANKL-induced osteoclastogenesis and might become a potential therapeutic drug for bone-resorptive diseases.

Modulation of SEMA4D-modified titanium surface on M2 macrophage polarization via activation of Rho/ROCK-mediated lactate release of endothelial cells: In vitro and in vivo.

SEMA4D-modified titanium surfaces can indirectly regulate macrophages through endothelial cells to achieve an anti-inflammatory effect, which is beneficial for healing soft tissues around the gingival abutment. However, the mechanism of surface-induced cellular phenotypic changes in SEMA4D-modified titanium has not yet been elucidated. SEMA4D activates the RhoA signaling pathway in endothelial cells, which coordinates metabolism and cytoskeletal remodeling. This study hypothesized that endothelial cells inoculated on SEMA4D-modified titanium surfaces can direct M2 polarization of macrophages via metabolites. An indirect co-culture model of endothelial cells and macrophages was constructed in vitro, and specific inhibitors were employed. Subsequently, endothelial cell adhesion and migration, metabolic changes, Rho/ROCK1 expression, and inflammatory expression of macrophages were assessed via immunofluorescence microscopy, specific kits, qRT-PCR, and Western blotting. Moreover, an in vivo rat bilateral maxillary implant model was constructed to evaluate the soft tissue healing effect. The in vitro experiments showed that the SEMA4D group had stronger endothelial cell adhesion and migration, increased Rho/ROCK1 expression, and enhanced release of lactate. Additionally, decreased macrophage inflammatory expression was observed. In contrast, the inhibitor group partially suppressed lactate metabolism and motility, whereas increased inflammatory expression. The in vivo analyses indicated that the SEMA4D group had faster and better angiogenic and anti-inflammatory effects, especially in the early stage. In conclusion, via the Rho/ROCK1 signaling pathway, the SEMA4D-modified titanium surface promotes endothelial cell adhesion and migration and lactic acid release, then the paracrine lactic acid promotes the polarization of macrophages to M2, thus obtaining the dual effects of angiogenesis and anti-inflammation.

Inhibition of Sema4D attenuates pressure overload-induced pathological myocardial hypertrophy via the MAPK/NF-κB/NLRP3 pathways.

Sema4D (CD100) is closely related to pathological and physiological processes, including tumor growth, angiogenesis and cardiac development. Nevertheless, the role and mechanism of Sema4D in cardiac hypertrophy are still unclear to date. To assess the impact of Sema4D on pathological cardiac hypertrophy, TAC surgery was performed on C57BL/6 mice which were transfected with AAV9-mSema4D-shRNA or AAV9-mSema4D adeno-associated virus by tail vein injection. Our results indicated that Sema4D knockdown mitigated cardiac hypertrophy, fibrosis and dysfunction when exposed to pressure overload, and Sema4D downregulation markedly inhibited cardiomyocyte hypertrophy induced by angiotensin II. Meanwhile, Sema4D overexpression had the opposite effect in vitro and in vivo. Furthermore, analysis of signaling pathways showed that Sema4D activated the MAPK pathway during cardiac hypertrophy induced by pressure overload, and the pharmacological mitogen-activated protein kinase kinase 1/2 inhibitor U0126 almost completely reversed Sema4D overexpression-induced deteriorated phenotype, resulting in improved cardiac function. Further research indicated that myocardial hypertrophy induced by Sema4D was closely related to the expression of the pyroptosis-related proteins PP65, NLRP3, caspase-1, ASC, GSDMD, IL-18 and IL-1ß. In conclusion, our study demonstrated that Sema4D regulated the process of pathological myocardial hypertrophy through modulating MAPK/NF-κB/NLRP3 pathway, and Sema4D may be the promising interventional target of cardiac hypertrophy and heart failure.

Cholangiocytes Modulate CD100 Expression in the Liver and Facilitate Pathogenic T-Helper 17 Cell Differentiation.

BACKGROUND & AIMS: Chronic inflammation surrounding bile ducts contributes to the disease pathogenesis of most cholangiopathies. Poor efficacy of immunosuppression in these conditions suggests biliary-specific pathologic principles. Here we performed biliary niche specific functional interpretation of a causal mutation (CD100 K849T) of primary sclerosing cholangitis (PSC) to understand related pathogenic mechanisms. METHODS: Biopsy specimens of explanted livers and endoscopy-guided sampling were used to assess the CD100 expression by spatial transcriptomics, immune imaging, and high-dimensional flow cytometry. To model pathogenic cholangiocyte-immune cell interaction, splenocytes from mutation-specific mice were cocultured with cholangiocytes. Pathogenic pathways were pinpointed by RNA sequencing analysis of cocultured cells and cross-validated in patient materials. RESULTS: CD100 is mainly expressed by immune cells in the liver and shows a unique pattern around PSC bile ducts with RNA-level colocalization but poor detection at the protein level. This appears to be due to CD100 cleavage as soluble CD100 is increased. Immunophenotyping suggests biliary-infiltrating T cells as the major source of soluble CD100, which is further supported by reduced surface CD100 on T cells and increased metalloproteinases in cholangiocytes after coculturing. Pathogenic T cells that adhered to cholangiocytes up-regulated genes in the T-helper 17 cell differentiation pathway, and the CD100 mutation boosted this process. Consistently, T-helper 17 cells dominate biliary-resident CD4 T cells in patients. CONCLUSIONS: CD100 exerts its functional impact through cholangiocyte-immune cell cross talk and underscores an active, proinflammatory role of cholangiocytes that can be relevant to novel treatment approaches.

Sema4D as a biomarker for Predicting rheumatoid arthritis disease activity.

OBJECTIVE: The semaphorins are membrane or secreted proteins first identified in neural development. Semaphorin 4D (Sema4D) is the first family member found to have immune properties. We evaluated the potential of Sema4D as a marker for rheumatoid arthritis (RA) disease activity, singly and in combination with other known biomarkers including rheumatoid factor (RF) and C-reactive protein (CRP). METHODS: Three hundred and eleven RA patients were enrolled. The patients were divided into three groups based on their disease activity in 28 joints (DAS28): mild, moderate, and severe. The healthy group included 40 healthy individuals. SerumSema4D was measured by quantitative ELISA and the specificity and sensitivity of biomarkers were evaluated by generating a receiver operating characteristic (ROC) curve to analyze their diagnostic accuracy. RESULTS: Serum Sema4D levels in the moderate and severe RA groups were elevated significantly above those of the controls (P < 0.01), while levels in the mild RA and control groups did not differ significantly (P > 0.05). The Sema4D cutoff threshold was 15.7 ng/ml when the DAS28 was applied as a reference. Compared to the erythrocyte sedimentation rate (ESR and CRP, Sema4D had the highest specificity (96.8%) and area under the curve (0.80) for diagnosing RA activity. The highest specificity (100%) for the biomarker combinations was obtained when Sema4D was combined with CRP and anti-CCP, the combination of the Sema4D combined with ESR and anti-CCP had the highest sensitivity (99.35%). According to this result, a new model for jointly calculating RA activity of Sema4D,anti-CCP and CRP was constructed. Meanwhile another model is established by using the method of multivariate analysis.Model comparison results showed the the multiple regression algorithm method fitted the patients' disease activity better. CONCLUSION: The serum Sema 4D level effectively reflects moderate to severe RA activity. Sema4D levels can be used together with conventional RA biomarkers to increase the diagnostic power of RA activity. The multiple regression algorithm method is promising in disease activity calculation.

Schistosome egg-derived extracellular vesicles deliver Sja-miR-71a inhibits host macrophage and neutrophil extracellular traps via targeting Sema4D.

BACKGROUND: Macrophages and neutrophils are rapidly recruited around Schistosome eggs to form granulomas. Extracellular traps (ETs) of macrophages and neutrophils are part of the pathogen clearance armamentarium of leukocytes. Schistosome eggs possess the ability to resist attack by the host's immune cells and survive by employing various immune evasion mechanisms, including the release of extracellular vesicles (EVs). However, the specific mechanisms by which Schistosome egg-derived EVs (E-EVs) evade the immune response and resist attack from macrophage and neutrophil ETs remain poorly understood. In this study, we aimed to investigate the association between E-EVs and macrophage/neutrophil ETs. METHODS: EVs were isolated from the culture supernatant of S. japonicum eggs and treated macrophages and neutrophils with E-EVs and Sja-miR-71a. The formation of ETs was then observed. Additionally, we infected mice with S. japonicum, administered HBAAV2/9-Sja-miR-71a, and the formation of macrophage ETs (METs) and neutrophil ETs (NETs) in the livers was measured. Sema4D-knockout mice, RNA sequencing, and trans-well assay were used to clarify Sja-miR-71a in E-EVs inhibits METs and NETs formation via the Sema4D/ PPAR-γ/ IL-10 axis. RESULTS: Our findings revealed that E-EVs were internalized by macrophages and neutrophils, leading to the inhibition of METs and NETs formation. The highly expressed Sja-miR-71a in E-EVs targeted Sema4D, resulting in the up-regulation of IL-10 and subsequent inhibition of METs and NETs formation. Sema4D knockout up-regulated IL-10 expression and inhibited the formation of METs and NETs. Furthermore, we further demonstrated that Sja-miR-71a inhibits METs and NETs formation via the Sema4D/ PPAR-γ/ IL-10 axis. CONCLUSIONS: In summary, our findings provide new insights into the immune evasion abilities of Schistosome eggs by demonstrating their ability to inhibit the formation of METs and NETs through the secretion of EVs. This study enhances our understanding of the host-pathogen interaction and may have implications for the development of novel therapeutic approaches. Video Abstract.

SEMA4D/VEGF surface enhances endothelialization by diminished-glycolysis-mediated M2-like macrophage polarization.

Cardiovascular disease remains the leading cause of death and morbidity worldwide. Inflammatory responses after percutaneous coronary intervention led to neoathrosclerosis and in-stent restenosis and thus increase the risk of adverse clinical outcomes. In this work, a metabolism reshaped surface is engineered, which combines the decreased glycolysis promoting, M2-like macrophage polarization, and rapid endothelialization property. Anionic heparin plays as a linker and mediates cationic SEMA4D and VEGF to graft electronically onto PLL surfaces. The system composed by anticoagulant heparin, immunoregulatory SEMA4D and angiogenic VEGF endows the scaffold with significant inhibition of platelets, fibrinogen and anti-thrombogenic properties, also noteworthy immunometabolism reprogram, anti-inflammation M2-like polarization and finally leading to rapid endothelializaiton performances. Our research indicates that the immunometabolism method can accurately reflect the immune state of modified surfaces. It is envisioned immunometabolism study will open an avenue to the surface engineering of vascular implants for better clinical outcomes.

Semaphorin 4D Induces Vasculogenic Differentiation of Dental Pulp Stem Cells.

This work aimed to evaluate the effect of Semaphorin 4D (SEMA4D) signaling through Plexin B1 on the vasculogenic differentiation of dental pulp stem cells. We assessed the protein expression of SEMA4D and Plexin B1 in dental pulp stem cells (DPSC) from permanent human teeth and stem cells from human exfoliated deciduous (SHED) teeth using Western blots. Their expression in human dental pulp tissues and DPSC-engineered dental pulps was determined using immunofluorescence. We then exposed dental pulp stem cells to recombinant human SEMA4D (rhSEMA4D), evaluated the expression of endothelial cell differentiation markers, and assessed the vasculogenic potential of rhSEMA4D using an in vitro sprouting assay. Lastly, Plexin B1 was silenced to ascertain its role in SEMA4D-mediated vasculogenic differentiation. We found that SEMA4D and Plexin B1 are expressed in DPSC, SHED, and human dental pulp tissues. rhSEMA4D (25-100 ng/mL) induced the expression of endothelial markers, i.e., vascular endothelial growth factor receptor (VEGFR)-2, cluster of differentiation (CD)-31, and tyrosine kinase with immunoglobulin-like and EGF-like domains (Tie)-2, in dental pulp stem cells and promoted capillary-like sprouting in vitro (p < 0.05). Furthermore, Plexin B1 silencing abrogated the vasculogenic differentiation of dental pulp stem cells and significantly inhibited capillary sprouting upon exposure to rhSEMA4D. Collectively, these data provide evidence that SEMA4D induces vasculogenic differentiation of dental pulp stem cells through Plexin B1 signaling.

Leptin and melatonin's effects on OVX rodents' bone metabolism.

Purpose: This study aims to examine the effects of leptin and melatonin intervention on bone metabolism in ovariectomize (OVX) rodents, as well as their potential mechanisms of action. Methods: Prepare an OVX model of osteoporosis in rodents and validate the model by collecting bilateral tibia samples for Micro-CT scanning and histological analysis. A control group of normal size, the OVX group, the OVX+Sema4D (Semaphorin 4D) group, the OVX+Sema4D+Leptin group, the OVX+Sema4D+ Melatonin(MT) group and the OVX+Sema4D+Leptin+ MT group were the experimental groups. Adenovirus vector construction and tibial medullary injection validation were conducted in accordance with the aforementioned experimental groups. Four groups of rats were injected with the Sema4D overexpression adenovirus vector into the tibial medullary cavity, and two groups were injected with the Leptin overexpression adenovirus vector. The repair of osteoporosis was observed using micro-CT and histological analysis. Immunohistochemical detection of bone morphogenetic protein-2 (BMP-2) expression in bone tissue was employed to ascertain the amount of osteoclasts in the upper tibial metaphysis, utilizing TRAP(tartrate-resistant acid phosphatase) staining. Results: Increased levels of BV/TV, Tb.N, BMD, and BMC were seen in the OVX+ Sema4D+Leptin, OVX+ Sema4D+MT, and OVX+ Sema4D+Leptin+ MT groups compared to the OVX group, whereas Tb. Sp levels were lowered. When compared to the Sema4D overexpression group, the trabecular bone structure of the OVX + Sema4D + Leptin, OVX + Sema4D + MT, and OVX + Sema4D + Leptin + MT groups is largely intact, tends to be closer, and the amount of trabecular bone increases. The OVX + Sema4D + Leptin + MT group in particular.The expression of BMP-2 was dramatically upregulated (p<0.05), the number of TRAP-stained osteoclasts was significantly reduced (p<0.05), and BALP(bone-derived alkaline phosphatase) and TRAP-5b(tartrate-resistant acid phosphatase-5b) activities were significantly downregulated (p<0.05). Conclusion: In rats with osteoporosis, leptin and melatonin can be seen to augment the trabecular microstructure of the bone, augment bone growth, diminish trabecular harm, and mend the bone. The combined effect is more powerful.

Mediating immunosuppressive functions: a new perspective on the complex immunological properties of SEMA4D in the tumor microenvironment.

Semaphorin 4D (SEMA4D) is considered a new antitumor target closely related to immune cells. However, understanding the role of SEMA4D in the tumor microenvironment (TME) is limited. In this study, we explored the expression and immune cell infiltration patterns of SEMA4D using multiple bioinformatics datasets and analyzed the relationship between SEMA4D expression with immune checkpoints, tumor mutational load (TMB), microsatellite instability (MSI) and immune function. We detected that SEMA4D is overexpressed in many tumors types, widely enriched in immune cells, and closely associated with TILs, MSI, TMB, as well as T-cell exhaustion-associated immune checkpoints, and thus can broadly affect the immune microenvironment. We further verified the overexpression of SEMA4D in tumor and its distribution in TME by immunohistochemistry, RT-qPCR and flow cytometry, and confirmed that decreased expression of SEMA4D can lead to recovery of T cell exhaustion. In conlusion, this study provides a more comprehensive perspective of SEMA4D regulation of tumor immunity, which provide a new option for cancer immunotherapy.

Semaphorin 4D induced inhibitory synaptogenesis decreases epileptiform activity and alters progression to Status Epilepticus in mice.

Previously we demonstrated that intra-hippocampal infusion of purified, Semaphorin 4D (Sema4D) extracellular domain (ECD) into the mouse hippocampus rapidly promotes formation of GABAergic synapses and decreases seizure susceptibility in mice. Given the relatively fast action of Sema4D treatment revealed by these studies, we sought to determine the time course of Sema4D treatment on hippocampal network activity using an acute hippocampal slice preparation. We performed long-term extracellular recordings from area CA1 encompassing a 2-hour application of Sema4D and found that hippocampal excitation is suppressed for hours following treatment. We also asked if Sema4D treatment could ameliorate seizures in an acute seizure model: the kainic acid (KA) mouse model. We demonstrate that Sema4D treatment delays and suppresses ictal activity, delays the transition to Status Epilepticus (SE), and lessens the severity of SE. Lastly, we sought to explore alternative methods of Sema4D delivery to hippocampus and thus created an Adeno Associated Virus expressing the ECD of Sema4D. Our data reveal that virally delivered, chronically overexpressed Sema4D-ECD promotes GABAergic synapse formation and suppresses ictal activity and progression to SE. These results provide proof of concept that viral delivery of Sema4D is an efficacious and promising delivery method to abate epileptiform activity and progression to SE.

Sema4D silencing increases the sensitivity of nivolumab to B16-F10 resistant melanoma via inhibiting the PI3K/AKT signaling pathway.

Melanoma is a common skin tumor that causes a high rate of mortality, especially in Europe, North America and Oceania. Immunosuppressants such as anti-PD-1 have been used in the treatment of malignant melanoma, however, nearly 60% of patients do not respond to these treatments. Sema4D, also called CD100, is expressed in T cells and tumor tissues. Sema4D and its receptor, Plexin-B1, play crucial roles in the process of immune regulation, angiogenesis, and tumor progression. The role of Sema4D in melanoma with anti-PD-1 resistance is poorly understood. Through a combination of molecular biology techniques and in silico analysis, the role of Sema4D in improving anti-PD-L1 sensitivity in melanoma was explored. The results showed that the expression of Sema4D, Plexin-B1 and PD-L1 was significantly increased in B16-F10R cells. Sema4D knockdown synergizes with anti-PD-1 treatment, cell viability, cell invasion and migration were significantly decreased, while the apoptosis was increased, the growth of tumors on the mice was also inhibited. Mechanistically, bioinformatics analysis revealed that Sema4D is involved in the PI3K/AKT signaling pathway; the downregulation of p-PI3K/PI3K and p-AKT/AKT expression were observed in Sema4D knockdown, therefore, nivolumab resistance is related to Sema4D and Sema4D silencing can improve sensitivity to nivolumab via inhibition of the PI3K/AKT signaling pathway.

Impact of leptin or melatonin on Sema4D overexpression-related bone metabolism.

PURPOSE: The current study aims to investigate the regulatory impact of leptin or melatonin on bone metabolism as well as the underlying mechanism in conjunction with Sema4D (monoclonal antibody to semaphorin 4D). METHODS: Rats were used to create the osteoporosis model utilizing the OVX (OVariectomize) technique. Rat tibial specimens from each side were collected for three-dimensional reconstruction and Micro-CT scanning examination. The Hematoxylin-osinstaining (HE) staining technique was used to determine the pathological condition of bone tissues. The ELISA (Enzyme-Linked Immunosorbent Assay) assay was used to measure the amount of estradiol present in the serum. In the current study, there were six groups: control, OVX, OVX + NL (no load group), OVX + Sema4D, OVX + Sema4D + leptin, and OVX + Sema4D + MT (melatonin). Rats were given injections of the Sema4D or leptin overexpressing vectors via the tail vein in accordance with the aforementioned classification. By using a high-resolution micro-CT technology, 3D bone structure was discovered. The activity of tartrate-resistant acid phosphatase-5b (TRAP-5b) and bone-derived alkaline phosphatase (BALP) in serum was assessed using an ELISA. The number of osteoclasts in the metaphysis of the upper tibia was determined using TRAP (tartrate-resistant acid phosphatase) staining. Immunohistochemistry was used to find leptin and bone morphogenetic protein-2 (BMP-2) expressions in bone tissue. RESULTS: The BV/TV (Bone volume/Tissue volume), Tb.N (Trabecular number), BMD (Bone Mineral Density), and BMC (Bone Mineral Content) levels were significantly higher in the OVX + Sema4D + leptin and OVX + Sema4D + MT groups compared to OVX + NL, while Tb.Sp (Trabecular separation) levels were significantly lower. In contrast to the OVX group, the bone trabeculae in the OVX + Sema4D + leptin and OVX + Sema4D + MT groups had a relatively complete structure and tended to be organized closely. The amount of bone trabeculae grew drastically, whereas the proportion of TRAP-positive osteoclasts declined dramatically. BMP-2 and leptin were also elevated, while BALP and TRAP-5b activity was reduced. CONCLUSION: Leptin or melatonin improved Sema4d's role in trabecular bone microstructure, bone production, and repairment of trabecular bone loss in osteoporosis rats.

Clinical values of serum Semaphorin 4D (Sema4D) in medication­related osteonecrosis of the jaw.

BACKGROUND: Bisphosphonates (BPs) are widely used in clinical practice to prevent and treat bone metabolism-related diseases. Medication-related osteonecrosis of the jaw (MRONJ) is one of the major sequelae of BPs use. Early prediction and intervention of MRONJ are of great significance. METHODS: Ninety-seven patients currently on treatment with BPs or with a history of BPs usage and 45 healthy volunteers undergoing dentoalveolar surgery were included in this study. Participants' serum Semaphorin 4D (Sema4D) levels were measured and analyzed before participants underwent surgery (T0) and after a 12-month follow-up (T1). Kruskal-Wallis test and ROC analysis were used to examine the predictive effect of Sema4D on MRONJ. RESULTS: Sema4D levels in serum of patients corresponding to confirmed MRONJ were significantly lower at both T0 and T1 time points compared to non-MRONJ and healthy controls. Sema4D has a statistically predictive effect on the occurrence and diagnosis of MRONJ. Serum Sema4D levels were significantly reduced in MRONJ class 3 patients. MRONJ patients who received intravenous BPs had significantly lower Sema4D levels than those who received oral BPs. CONCLUSION: Serum Sema4D level has predictive value for the onset of MRONJ in BPs users within 12 weeks after dentoalveolar surgery.

SEMA4D/PlexinB1 promotes AML progression via activation of PI3K/Akt signaling.

BACKGROUND: Acute myeloid leukemia (AML) is the most common type of acute leukemia in adults. SEMA4D is a 150 kDa transmembrane protein that belongs to the IV class of the subfamily of semaphorin family. Previous studies have reported that SEMA4D is a multifunctional target in many solid tumors, involving multiple physiological systems, and there are emerging therapies to target these pathways. The role of SEMA4D in AML has not yet been explored. METHODS: The SEMA4D expression prolile, clinical data and potential prognostic analysis were acquired via the cBioPortal and GEPIA databases. SEMA4D expression was measured using real-time quantitative PCR and western blot. Cell counting kit-8 (CCK8) and flow cytometry were used to evaluate the malignant biological characteristics. RESULTS: We observed that SEMA4D was increased in AML patients and correlated with risk stratification and prognosis. Moreover, SEMA4D promotes the proliferation and inhibits apoptosis of AML cells by binding to its receptor, PlexinB1, and reduces the sensitivity of AML cells to daunorubicin. In addition, SEMA4D/PlexinB1 promotes the proliferation and survival of AML cells by activating the PI3K/Akt signaling pathway. VX15/2503, an anti-SEMA4D antibody, can inhibit the proliferation of AML cells in xenograft mouse models, thereby inhibiting the development of AML. CONCLUSION: SEMA4D will serve as a unique predictive biomarker and a possible therapeutic target in AML.

An antagonistic monoclonal anti-Plexin-B1 antibody exerts therapeutic effects in mouse models of postmenopausal osteoporosis and multiple sclerosis.

Osteoporosis and multiple sclerosis are highly prevalent diseases with limited treatment options. In light of these unmet medical needs, novel therapeutic approaches are urgently sought. Previously, the activation of the transmembrane receptor Plexin-B1 by its ligand semaphorin 4D (Sema4D) has been shown to suppress bone formation and promote neuroinflammation in mice. However, it is unclear whether inhibition of this receptor-ligand interaction by an anti-Plexin-B1 antibody could represent a viable strategy against diseases related to these processes. Here, we raised and systematically characterized a monoclonal antibody directed against the extracellular domain of human Plexin-B1, which specifically blocks the binding of Sema4D to Plexin-B1. In vitro, we show that this antibody inhibits the suppressive effects of Sema4D on human osteoblast differentiation and mineralization. To test the therapeutic potential of the antibody in vivo, we generated a humanized mouse line, which expresses transgenic human Plexin-B1 instead of endogenous murine Plexin-B1. Employing these mice, we demonstrate that the anti-Plexin-B1 antibody exhibits beneficial effects in mouse models of postmenopausal osteoporosis and multiple sclerosis in vivo. In summary, our data identify an anti-Plexin-B1 antibody as a potential therapeutic agent for the treatment of osteoporosis and multiple sclerosis.