Inflammation in the Peripheral Nervous System after Injury.

Nerve injury is a common condition that occurs as a result of trauma, iatrogenic injury, or long-lasting stimulation. Unlike the central nervous system (CNS), the peripheral nervous system (PNS) has a strong capacity for self-repair and regeneration. Peripheral nerve injury results in the degeneration of distal axons and myelin sheaths. Macrophages and Schwann cells (SCs) can phagocytose damaged cells. Wallerian degeneration (WD) makes the whole axon structure degenerate, creating a favorable regenerative environment for new axons. After nerve injury, macrophages, neutrophils and other cells are mobilized and recruited to the injury site to phagocytose necrotic cells and myelin debris. Pro-inflammatory and anti-inflammatory factors involved in the inflammatory response provide a favorable microenvironment for peripheral nerve regeneration and regulate the effects of inflammation on the body through relevant signaling pathways. Previously, inflammation was thought to be detrimental to the body, but further research has shown that appropriate inflammation promotes nerve regeneration, axon regeneration, and myelin formation. On the contrary, excessive inflammation can cause nerve tissue damage and pathological changes, and even lead to neurological diseases. Therefore, after nerve injury, various cells in the body interact with cytokines and chemokines to promote peripheral nerve repair and regeneration by inhibiting the negative effects of inflammation and harnessing the positive effects of inflammation in specific ways and at specific times. Understanding the interaction between neuroinflammation and nerve regeneration provides several therapeutic ideas to improve the inflammatory microenvironment and promote nerve regeneration.

Interactions between Schwann cell and extracellular matrix in peripheral nerve regeneration.

Peripheral nerve injuries, caused by various reasons, often lead to severe sensory, motor, and autonomic dysfunction or permanent disability, posing a challenging problem in regenerative medicine. Autologous nerve transplantation has been the gold standard in traditional treatments but faces numerous limitations and risk factors, such as donor area denervation, increased surgical complications, and diameter or nerve bundle mismatches. The extracellular matrix (ECM) is a complex molecular network synthesized and released into the extracellular space by cells residing in tissues or organs. Its main components include collagen, proteoglycans/glycosaminoglycans, elastin, laminin, fibronectin, etc., providing structural and biochemical support to surrounding cells, crucial for cell survival and growth. Schwann cells, as the primary glial cells in the peripheral nervous system, play various important roles. Schwann cell transplantation is considered the gold standard in cell therapy for peripheral nerve injuries, making ECM derived from Schwann cells one of the most suitable biomaterials for peripheral nerve repair. To better understand the mechanisms of Schwann cells and the ECM in peripheral nerve regeneration and their optimal application, this review provides an overview of their roles in peripheral nerve regeneration.

Fas ligand regulate nerve injury and repair by affecting AKT, ß-catenin, and NF-κB pathways.

Objective: To investigate the regulatory effect of Fas-L on the repair and regeneration of peripheral extension injury in rats. Methods: This study aimed to explore the effects of Fas-L on apoptosis and axonal regeneration of dorsal root ganglion (DRG) cells in rat peripheral nerve repair and regeneration by using several relevant experimental techniques from the injured nerve animal model, cell biology, and molecular biology. Results: The expression level of Fas-L in DRG tissues was significantly down-regulated after sciatic nerve injury. Interference with Fas-L can significantly promote the regeneration of DRG neuronal axons and inhibit apoptosis, while the overexpression of Fas-L is contrary to it. Moreover, Fas-L may play a role in the regulation of DRG function and the repair and regeneration of peripheral nerves in Sprague Dawley (SD) rats by affecting several signaling pathways, such as p-AKT/AKT, ß-catenin, and NF-κB. Conclusion: Fas-L may have a certain effect on the repair and regeneration of peripheral nerve injury in SD rats, which may provide an experimental basis and a new theoretical basis for the functional reconstruction of peripheral nerves. Significance statement: The expression level of Fas-L in DRG tissues was significantly down-regulated after sciatic nerve injury. Fas-L can significantly promote the regeneration of DRG neuronal axons and inhibit apoptosis. Fas-L may play a role in the regulation of DRG function and the repair and regeneration of peripheral nerves in SD rats by affecting several signaling pathways, such as p-AKT/AKT, ß-catenin, and NF-κB. Fas-L may have a certain effect on the repair and regeneration of peripheral nerve injury in SD rats, which may provide an experimental basis and a new theoretical basis for the functional reconstruction of peripheral nerves.

Role of Long Non-coding RNA in Nerve Regeneration.

Nerve injury can be caused by a variety of factors. It often takes a long time to repair a nerve injury and severe nerve injury is even difficult to heal. Therefore, increasing attention has focused on nerve injury and repair. Long non-coding RNA (lncRNA) is a newly discovered non-coding RNA with a wide range of biological activities. Numerous studies have shown that a variety of lncRNAs undergo changes in expression after nerve injury, indicating that lncRNAs may be involved in various biological processes of nerve repair and regeneration. Herein, we summarize the biological roles of lncRNAs in neurons, glial cells and other cells during nerve injury and regeneration, which will help lncRNAs to be better applied in nerve injury and regeneration in the future.

Long noncoding RNA H19 regulates degeneration and regeneration of injured peripheral nerves.

Our previous studies have shown that long noncoding RNA (lncRNA) H19 is upregulated in injured rat sciatic nerve during the process of Wallerian degeneration, and that it promotes the migration of Schwann cells and slows down the growth of dorsal root ganglion axons. However, the mechanism by which lncRNA H19 regulates neural repair and regeneration after peripheral nerve injury remains unclear. In this study, we established a Sprague-Dawley rat model of sciatic nerve transection injury. We performed in situ hybridization and found that at 4-7 days after sciatic nerve injury, lncRNA H19 was highly expressed. At 14 days before injury, adeno-associated virus was intrathecally injected into the L4-L5 foramina to disrupt or overexpress lncRNA H19. After overexpression of lncRNA H19, the growth of newly formed axons from the sciatic nerve was inhibited, whereas myelination was enhanced. Then, we performed gait analysis and thermal pain analysis to evaluate rat behavior. We found that lncRNA H19 overexpression delayed the recovery of rat behavior function, whereas interfering with lncRNA H19 expression improved functional recovery. Finally, we examined the expression of lncRNA H19 downstream target SEMA6D, and found that after lncRNA H19 overexpression, the SEMA6D protein level was increased. These findings suggest that lncRNA H19 regulates peripheral nerve degeneration and regeneration through activating SEMA6D in injured nerves. This provides a new clue to understand the role of lncRNA H19 in peripheral nerve degeneration and regeneration.

Interleukin-35 attenuates blood-brain barrier dysfunction caused by cerebral ischemia-reperfusion injury through inhibiting brain endothelial cell injury.

Background: Interleukin-35 (IL-35), an anti-inflammatory and antioxidant cytokine, plays a potent immunosuppressive role in various diseases. However, the effects of IL-35 on blood-brain barrier (BBB) dysfunction in ischemic stroke are not well characterized. Methods: A total of 150 male C57BL/6 mice (aged 6-8 weeks and weighing 20-25 g) were used in this study. The protective effects of IL-35 against BBB dysfunction were examined using a mouse model of middle cerebral artery occlusion (MCAO) and an in vitro model of oxygen-glucose deprivation/reoxygenation (OGD/R) injury in mouse brain endothelial cells (bEnd.3). Results: Intracerebroventricular administration of IL-35 (10 µg/g) was found to reduce cerebral edema and Evans blue (EB) leakage, and increase the expression of tight junction (TJ) proteins, thereby attenuating MCAO-induced neurological deficit in mice. Moreover, IL-35 (20 ng/mL) treatment upregulated the expression of TJ proteins in OGD/R-induced bEnd.3 cells. IL-35 also markedly suppressed the expression of caspase-1, IL-1ß, and gasdermin D (GSDMD) in vivo and in vitro. In addition, IL-35 decreased the generation of reactive oxygen species (ROS) and inhibited the expression of thioredoxin-interacting protein (TXNIP) in OGD/R-induced bEnd.3 cells. Conclusions: These results indicated that IL-35 exerts a protective effect on the BBB by targeting the ROS/TXNIP/caspase-1 pathway in cerebral ischemia-reperfusion (I/R) injury.

CtBP2 interacts with TGIF to promote the progression of esophageal squamous cell cancer through the Wnt/ß­catenin pathway.

C­terminal­binding protein 2 (CtBP2), a transcriptional co­repressor, plays a main role in tumorigenesis and in the development of multiple tumors. Transforming growth interacting factor (TGIF) is involved in a number of cellular signal transduction pathways and is related to tumor occurrence and development. In the present study, the proteins interacting with CtBP2 were identified and the mechanisms underlying the biological activity of CtBP2 in esophageal squamous cell carcinoma (ESCC) were investigated. The Search Tool for the Retrieval of Interacting Genes (STRING) database was used to search for known proteins interacting with CtBP2, and co­immunoprecipitation (Co­IP) assay was performed to validate the interactions. Reverse transcription­quantitative PCR (RT­qPCR), immunohistochemistry (IHC) and western blot analysis were performed to examine the expression levels of CtBP2 and TGIF in ESCC. The correlation between CtBP2 and TGIF was analyzed using Gene Expression Profiling Interactive Analysis (GEPIA) by Pearson's correlation analysis, and the co­localization of CtBP2 with TGIF in the ECA109 cells was identified using immunofluorescence staining. XAV939 treatment, CCK­8, 5­ethynyl­2'­deoxyuridine (EdU) staining, wound healing and Transwell assays were performed to investigate the signaling pathways involved in the biological activity of CtBP2 in ECA109 cells. According to the results obtained from STRING and Co­IP analysis, an interaction between CtBP2 and TGIF was indicated, and these proteins were co­localized in the nucleus. CtBP2 and TGIF mRNA and protein expression levels were robustly and simultaneously increased in both ESCC tissues and cell lines. There was a direct correlation between CtBP2 and TGIF expression levels in ESCC tissues, and both were significantly associated with metastasis and survival. The TGIF and CtBP2 expression levels were significantly increased or decreased simultaneously, in ECA109 cells transfected with LV­CtBP2 or sh­CtBP2, and vice versa. According to the results of CCK­8 assay, EdU staining and Transwell assay, CtBP2 promoted the proliferation, migration and invasion of ECA109 cells through the Wnt/ß­catenin pathway. On the whole, the present study demonstrates that CtBP2 interacts with TGIF and promotes the malignant progression of ESCC through the Wnt/ß­catenin pathway.

Characterization of rat and mouse acidic milk oligosaccharides based on hydrophilic interaction chromatography coupled with electrospray tandem mass spectrometry.

Oligosaccharides are one of the most important components in mammalian milk. Milk oligosaccharides can promote colonization of gut microbiota and protect newborns from infections. The diversity and structures of MOs differ among mammalian species. MOs in human and farm animals have been well-documented. However, the knowledge on MOs in rat and mouse have been very limited even though they are the most-widely used models for studies of human physiology and disease. Herein, we use a high-sensitivity online solid-phase extraction and HILIC coupled with electrospray tandem mass spectrometry to analyze the acidic MOs in rat and mouse. Among the fifteen MOs identified, twelve were reported for the first time in rat and mouse together with two novel sulphated oligosaccharides. The complete list of acidic oligosaccharides present in rat and mouse milk is the baseline information of these animals and should contribute to biological/biomedical studies using rats and mice as models.

Cardiac arrest and resuscitation activates the hypothalamic-pituitary-adrenal axis and results in severe immunosuppression.

In patients who are successfully resuscitated after initial cardiac arrest (CA), mortality and morbidity rates are high, due to ischemia/reperfusion injury to the whole body including the nervous and immune systems. How the interactions between these two critical systems contribute to post-CA outcome remains largely unknown. Using a mouse model of CA and cardiopulmonary resuscitation (CA/CPR), we demonstrate that CA/CPR induced neuroinflammation in the brain, in particular, a marked increase in pro-inflammatory cytokines, which subsequently activated the hypothalamic-pituitary-adrenal (HPA) axis. Importantly, this activation was associated with a severe immunosuppression phenotype after CA. The phenotype was characterized by a striking reduction in size of lymphoid organs accompanied by a massive loss of immune cells and reduced immune function of splenic lymphocytes. The mechanistic link between post-CA immunosuppression and the HPA axis was substantiated, as we discovered that glucocorticoid treatment, which mimics effects of the activated HPA axis, exacerbated post-CA immunosuppression, while RU486 treatment, which suppresses its effects, significantly mitigated lymphopenia and lymphoid organ atrophy and improved CA outcome. Taken together, targeting the HPA axis could be a viable immunomodulatory therapeutic to preserve immune homeostasis after CA/CPR and thus improve prognosis of post-resuscitation CA patients.

Development of chimeric antigen receptor-modified T cells for the treatment of esophageal cancer.

BACKGROUND: Human epidermal growth factor receptor 2 (HER2) is an overexpressed antigen in esophageal squamous cell carcinomas (ESCCs) but with limited expression levels in normal esophageal tissues. Therefore, employing the adoptive transfer of T cells genetically modified to express chimeric antigen receptor (CAR) targeting HER2 could be a promising therapeutic strategy against ESCC. METHODS: Two different second-generation CAR-T cells expressing antibodies for HER2 and CD19 antigens were developed using retroviral vector transduction. The expression of HER2 antigen in ESCC tissue and cell lines was examined by immunohistochemistry and flow cytometry, respectively. The tumor killing efficacy of the CAR-T cells in mice model and ESCC cell lines and its potential for the treatment of ESCC was evaluated by determining tumor size in mice xenograft, and by crystal violet staining, MTS assay, and cytokine release. RESULTS: In vitro, HER2.CAR-T cells efficiently recognized and killed HER2-positive tumor cells as evidenced by the secretion of proinflammatory cytokines, interferon-γ, and interleukin 2 and by cytotoxicity assays. In vivo, intratumor injection of HER2.CAR-T cells resulted in a significant suppression of established ESCCs in a subcutaneous xenograft BALB/c nude mouse model. In contrast, the injection of CD19.CAR-T cells did not affect the tumor growth pattern. CONCLUSIONS: An effective HER2 CAR targeting ESCC was developed successfully. The HER2.CAR-T cell showed promising immunotherapeutic potential for the treatment of HER2-positive esophageal cancer.

MCC950, a selective NLPR3 inflammasome inhibitor, improves neurologic function and survival after cardiac arrest and resuscitation.

BACKGROUND: Cardiac arrest (CA) is associated with high morbidity and mortality, even after spontaneous circulation is re-established. This dire situation is partly due to post-CA syndrome for which no specific and effective intervention is available. One key component of post-CA syndrome is sterile inflammation, which affects various organs including the brain. A major effector of sterile inflammation is activated NLRP3 inflammasome, which leads to increased release of interleukin (IL)-1ß. However, how NLRP3 inflammasome impacts neuroinflammation and neurologic outcome after CA is largely undefined. METHODS: Mice were subjected to a potassium-based murine CA and cardiopulmonary resuscitation (CPR) model. MCC950 was used to suppress activation of NLRP3 inflammasome after CA/CPR. Levels of protein and mRNA were examined by Western blotting and quantitative PCR, respectively. Immunologic changes were assessed by measuring cytokine expression and immune cell compositions. CA outcomes, including neurologic deficits, bacterial load in the lung, and survival rate, were evaluated. RESULTS: Using our CA/CPR model, we found that NLRP3 inflammasome was activated in the post-CA brain, and that pro-inflammatory cytokine levels, including IL-1ß, were increased. After treatment with MCC950, a potent and selective NLRP3 inflammasome inhibitor, mice exhibited improved functional recovery and survival rate during the 14-day observational period after CA/CPR. In line with these findings, IL-1ß mRNA levels in the post-CA brain were significantly suppressed after MCC950 treatment. Interestingly, we also found that in MCC950- vs. vehicle-treated CA mice, immune homeostasis in the spleen was better preserved and bacterial load in the lung was significantly reduced. CONCLUSIONS: Our data demonstrate that activation of NLRP3 inflammasome could be a key event shaping the post-CA immuno- and neuro-pathology, and identify this pathway as a unique and promising therapeutic target to improve outcomes after CA/CPR.

miR-203a-3p regulates the cellular processes of esophageal cancer cells via targeting CtBP2.

BACKGROUND: MicroRNAs (miRNA) (small noncoding RNAs) are vital modulators of gene expression by mRNA degradation and translational silencing. However, the definite mechanism and character of miR-203a-3p in regulating esophageal carcinoma cells remain unexplained. Here we further investigate the effect and the latent target gene of miR-203a-3p on the progression of esophageal squamous cell cancer (ESCC) tissues and cells. METHODS: The expressions of miR-203a-3p in ESCC tissues and peri-neoplastic tissues were further measured by RT-quantitative-PCR (RT-qPCR). Luciferase assay was applied to confirm that C-terminal-binding protein 2 (CtBP2) was the potential target gene of miR-203a-3p. miRNA mimic was transfected into ECA109 cells to up-regulate the miR-203a-3p expression, and its and CtBP2 expression were tested using RT-qPCR and Western blot. In vitro, MTT, transwell, wound healing, TUNEL and flow cytometry (FCM) assay were used to explore the role of miR-203a-3p on the cellular processes of ECA109 cells via targeting CtBP2. Furthermore, we designed rescue experiments by using CtBP2 stable over-expression ECA109 cells. RESULTS: We found the miR-203a-3p expressions in ESCC tissues and cells were significantly raised. miR-203a-3p negatively regulated the CtBP2 expression, and caused to inhibiting proliferation, migration and invasion, and promoting apoptosis in ECA109 cell. In addition, proteins involved in epithelial-mesenchymal transition (EMT) were measured by Western blot in ECA109 cells. miR-203a-3p enhanced the E-cadherin and ß-catenin expression, while reduced vimentin expression in ECA109 cells. In vivo, Xenograft tumor model demonstrated that tumor volume in miR-203a-3p agomir group was remarkably decreased. CONCLUSIONS: miR-203a-3p plays a vital role in the metastasis of ESCC cell by targeting CtBP2, and offers a promising therapeutic target for ESCC treatment.

Long non-coding RNA DANCR promotes cell proliferation, migration, invasion and resistance to apoptosis in esophageal cancer.

BACKGROUND: Long non-coding RNAs (lncRNAs) have important effects on the development and progression of multiple carcinomas. Our studies aimed to investigate the expression of lncRNA DANCR in esophageal squamous cell carcinoma (ESCC) tissues and paracancerous tissues, and to explore its effect on the cell biological characteristics of ESCC ECA109 cells. METHODS: The expression of DANCR was detected by qRT-PCR in human ESCC tissues and paracancerous normal tissues in ESCC patients. Small interfering RNA (siRNA) was transfected to knock down the expression of DANCR and interference efficiency was analyzed by qRT-PCR in ECA109 cells. MTT, wound healing, Transwell, TUNEL and flow cytometry (FCM) assay was used to measure the influence of DANCR on proliferation, invasion, migration and apoptosis in ECA109 cells, respectively. RESULTS: The expression of DANCR in ESCC tissues and ESCC cells was significantly higher compared with that in the adjacent normal tissues (P<0.05). Furthermore, cell proliferation, migration and invasion were significantly suppressed by knock-down mediated down-regulation of DANCR expression. On the contrary, cell apoptosis was promoted by silencing of DANCR. CONCLUSIONS: According to our research, the expression of DANCR was up-regulated in human ESCC tissues, and the important role that DANCR played in ESCC cells was similar to an oncogene. Therefore, silencing of lncRNA DANCR could have potentially beneficial effects on the prognostic and therapy for ESCC in the future.

EphA2 chimeric antigen receptor-modified T cells for the immunotherapy of esophageal squamous cell carcinoma.

BACKGROUND: It is urgent to explore an effective potential therapeutic strategy for ESCC. In recent years, cell-based cancer immunotherapy has become a potentially close for carcinoma therapy. Chimeric antigen receptor (CAR) T cell technology is a kind of adoptive cell therapy technique which has been developed rapidly. We sought to obtain EphA2.CAR-T cell and revealed the ability of EphA2.CAR-T cells to kill esophageal squamous cell carcinoma (ESCC) cells in vitro. METHODS: Firstly, the expression and location of EphA2 in ESCC tissues and cells was tested by immunohistochemistry staining and Western blot. Secondly, the second generation of EphA2.CAR was constructed via molecular biology technology, and transduced into T cells to obtain the EphA2.CAR-T cell. The transduction efficacies were assessed using flow cytometry (FCM). Thirdly, the effect of cell killing of EphA2.CAR-T cell on ESCC cells in vitro was detected by co-culture experiments. The productions of cytokines (TNF-α and IFN-γ) by EphA2.CAR-T cell after co-culture with ESCC cells were analyzed by ELISA assay. RESULTS: The expression of EphA2 was significantly upregulated in ESCC tissues and cells (P<0.05). EphA2 was expressed on the membrane of ESCC cells, so it could be served as tumor-associated surface antigens (TAA) of CAR for ESCC treatment. The EphA2.CAR-T cell was obtained successfully, and its' transduction efficacies was 61.4% by FCM. The ability of cell killing of EphA2.CAR-T cell was better than that of T cells (P<0.01), and demonstrated a dose-dependent cell killing. The results of ELISA assay showed that the levels of TNF-α and IFN-γ in EphA2.CAR-T cells were notably raised compared with T cells (P<0.05). CONCLUSIONS: We firstly constructed the second generation of EphA2.CAR and established EphA2.CAR-T cells. The EphA2.CAR-T cells showed a dose-dependent cell killing of ESCC cells, and promoted the production of cytokines in vitro. These findings open a new way for treatment of ESCC by immunotherapy in the future.

C-terminal binding protein­2 mediates cisplatin chemoresistance in esophageal cancer cells via the inhibition of apoptosis.

C-terminal binding protein­2 (CtBP2) is a transcriptional co-repressor that is associated with tumorigenesis and tumor progression. It has been reported to predict a poor prognosis in several human cancers, including esophageal squamous cell carcinoma (ESCC). The present study aimed to investigate the involvement of CtBP2 in the cisplatin (DDP) resistance of the ECA109 ESCC cell line and its effect on the expression of apoptosis-associated proteins. Constructed recombinant lentiviruses were used for the knockdown or overexpression of CtBP2 in ECA109 cells, and the expression of CtBP2 was measured using reverse transcription-quantitative polymerase chain reaction and western blotting following transfection. MTT assays, Hoechst 33342 staining and flow cytometry (FCM) were applied to detect the influence of CtBP2 on the DDP-induced viability and apoptosis of the transfected ECA109 cells. In addition, the levels of apoptosis-associated proteins, including p53, B­cell lymphoma 2 (Bcl­2), Bcl­2­associated X protein (Bax) and activated caspase-3 were investigated in the transfected ECA109 cells. Stable ECA109 cells with CtBP2 overexpression or knockdown were successfully established. The results of the MTT, Hoechst 33342 and FCM assays demonstrated that overexpression of CtBP2 attenuated the reduction of cell viability and inhibited the cell apoptosis induced by DDP. Furthermore, the western blotting results indicated that CtBP2 overexpression inhibited the DDP-induced apoptosis of ECA109 cells via the reduction of p53, activated caspase-3 and Bax expression, and promotion of Bcl­2 expression. Therefore, the present study indicated that CtBP2 reduced the susceptibility of ECA109 cells to DDP by regulating the expression of apoptosis-related proteins, suggesting that it may be a promising therapeutic target in ESCC in the future.

Correlation between sex and efficacy of immune checkpoint inhibitors (PD-1 and CTLA-4 inhibitors).

Immune checkpoint inhibitors (ICIs) exert the antitumor efficacy depending on immune response, which is affected by sex difference, where both biological and sociological factors are involved. The role of sex in ICI trials has been overlooked. How sex correlates with ICI efficacy is incompletely understood. Clinical trials evaluating ICI versus other therapies in male and female patients were included. The hazard ratio (HR) and 95% confidence interval (CI) of overall survival (OS) and progression-free survival (PFS) were used. Six thousand and ninety-six patients from 11 trials were included. More improvement of OS was observed in males (HR, 0.62; 95% CI, 0.53-0.71; p < 0.001) treated with ICI versus controls than females (HR, 0.74; 95% CI, 0.65-0.84; p < 0.001). ICIs improved PFS more in males (HR, 0.57; 95% CI, 0.43-0.71; p < 0.001) than females (HR, 0.71; 95% CI, 0.52-0.91; p < 0.001). The sex difference had more effect on the overall survival in melanoma patients versus NSCLC patients. Overall survival of patients treated with CTLA-4 inhibitor was more influenced by sex variable compared with PD-1 inhibitors. A significant sex-related efficacy difference was observed between female and male melanoma patients. Although male patients had longer OS and PFS than females when treated with ICIs versus controls, the difference was not significant. Sex difference should be more considered in future clinical trials, guidelines and clinical practice.

The clinical value of combination of immune checkpoint inhibitors in cancer patients: A meta-analysis of efficacy and safety.

The use of immune checkpoint inhibitors (ICIs) in combination therapy is an emerging trend in tumor immunology. However, the value of combination immunotherapy remains controversial, because of the toxic effects induced by combination. The added benefit of each additional drug has not been assessed against the added toxicity. We searched for clinical trials that evaluated ICI monotherapies and combination therapies in lung cancer and melanoma patients. The overall response rate (ORR), grade 3/4 treatment-related adverse event rate, overall survival (OS), and progression-free survival (PFS) were extracted from the most recently published studies to determine the relative risk (RR), hazard ratios (HRs), and 95% confidence intervals (CIs). Seven randomized controlled trials and one open-label study were identified (n = 3,097). Treatments included combinations of several ICIs, a combination of an ICI and dacarbazine, two combinations of an ICI, paclitaxel and carboplatin, and a combination of an ICI and gp100 vaccine. Higher ORR (RR: 1.51, 95% CI: 1.03-2.20, p = 0.034), OS (HR: 0.86, 95% CI: 0.78-0.95, p = 0.000), and PFS (HR: 0.93, 95% CI: 0.72-1.14, p = 0.000) values were observed in combination therapy than in monotherapy. In addition, the toxicity of combination ICI immunotherapy was higher (RR: 1.50, 95% CI: 1.03-2.19, p = 0.036) than that of monotherapy. This meta-analysis showed that the addition of nivolumab to ipilimumab better benefits PFS and ORR. Adding sargramostim was associated with better OS and safety. The efficacy and safety of a nivolumab-ipilimumab-sargramostim combination should be investigated further.

Syntaxin-4 and SNAP23 act as exocytic SNAREs to release NGF from cultured Schwann cells.

Nowadays peripheral nerve (PN) injury occurs more frequently, the outcome is often poor because of the ineffective treatment. Once the PN was injured, Schwann cells (SCs) release neurotrophins to guide the regeneration of axons. Recent researches revealed the duration of NGF administration acts a positive role during the nerve regeneration, but the molecular mechanisms of NGF release from SCs are unknown. To investigate components of the exocytic machinery of NGF, we used RT-PCR, Western blot and immunocytochemistry to investigate expressions and locations of soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs) in rat primary cultured SCs. We found that Syntaxin-4 and SNAP23 were co-localized with NGF by immunocytochemistry. Co-immunoprecipitation (Co-IP) and RNA interference (RNAi) confirmed Syntaxin-4 associated with SNAP23 to regulate the release of NGF from SCs. Knockdown of Syntaxin-4 and SNAP23 dramatically decreased the exocytosis of NGF and inhibited the neurite outgrowth of dorsal root ganglia (DRG). Syntaxin-4 and SNAP23 acted as exocytic SNAREs to release NGF from SCs.

The beneficial effect of chitooligosaccharides on cell behavior and function of primary Schwann cells is accompanied by up-regulation of adhesion proteins and neurotrophins.

Chitosan-based tissue engineered nerve grafts are successfully used for bridging peripheral nerve gaps. The biodegradation products of chitosan are water-dissolvable chitooligosaccharides (COSs), which have been shown to support peripheral nerve regeneration. In this study, we aimed to examine in vitro interactions between COSs and Schwann cells (SCs), the principal glial cells in the peripheral nervous system. Treatment of primary SCs with COSs enhanced cell survival and promoted cell proliferation in a dose-dependent manner (0.25-1.0 mg/ml), as determined by real-time cell analyzer-based assay, cell growth assay, cell cycle analysis, and EdU incorporation. Western blot analysis and immunocytochemistry with antibodies against MBP and MAG (two myelin-specific markers) showed that COSs enhanced axonal myelination in a co-culture system consisting of SCs and dorsal root ganglia (DRGs). Furthermore, we observed that COSs enhanced the protein expression of N-cadherin and ß-catenin in primary SCs, and also increased the release of BDNF and NGF in co-culture of SCs with DRGs. And we also noted that knockdown of N-cadherin in primary SCs reduced COSs-induced increase in cell proliferation. Our findings suggested that beneficial effects of COSs on cell behavior and functions of primary SCs might be accompanied by up-regulation of adhesion proteins and neurotrophins, thus providing a new insight into the supportive role of COSs during peripheral nerve regeneration.

Neural activity analysis of pure chito-oligomer components separated from a mixture of chitooligosaccharides.

Chitooligosaccharides (COSs) are obtained from the in vivo and in vitro degradation of chitosan, which is a natural biomaterial used successfully for peripheral nerve repair. Also, COSs have been reported to be beneficial for the nervous system and especially able to promote peripheral nerve regeneration. To determine which component in a mixture of COSs was really responsible for the neurotrophic action of COSs, this study was performed to separate the active components of COSs and compare the neural activity of different pure components. Hydrophilic interaction liquid chromatography (HILIC) was used to separate 5 chito-oligomers with single degrees of polymerization (DPs) from a mixture of COSs. They were chitobiose, chitotriose, chitotetraose, chitopentaose, and chitohexaose with DPs of 2-6, respectively. MTT assay indicated that chitotriose induced the greatest increase in Schwann cell survival among 5 chito-oligomers. Immunocytochemistry with anti-NF-200 showed that chitotriose significantly encouraged neurite outgrowth from dorsal root ganglion (DRG) explants with the greatest effect among 5 chito-oligomers. Our results suggest that chitotriose may be a key component in COSs mixture.