Synthesis of Fluorine-Containing Multisubstituted Oxa-Spiro[4,5]cyclohexadienones via a Fluorinated Alcohol-Catalyzed One-Pot Sequential Cascade Strategy.

In recent years, the application of fluorinated alcohols as solvents, cosolvents, or additives has become important in modern organic synthesis. However, their potential as efficient catalysts in organic synthesis has not been well-explored. In this article, we report on the development of a one-pot sequential cascade reaction of p-quinone methides with difluoroenoxysilanes using hexafluoroisopropanol as catalyst. This reaction allows for the preparation of fluorinated multisubstituted oxa-spiro[4,5]cyclohexadienones. By using 50 mol % 1,1,1,3,3,3-Hexafluoroisopropanol (HFIP), the reaction proceeds smoothly to yield 1,6-conjugated products, which are then subjected to oxidative dearomatization/hemiacetalization using PhI(OAc)2. The overall process affords moderate to high yields and excellent diastereoselectivities.

Palladium-Catalyzed C-H Allylation/Annulation Reaction of Amides and Allylic Alcohols: Regioselective Construction of Vinyl-Substituted 3,4-Dihydroisoquinolones.

A palladium-catalyzed highly regioselective C-H allylation/annulation reaction of N-sulfonyl amides with secondary or tertiary allylic alcohols has been developed to construct 3,4-dihydroisoquinolones bearing a synthetically valuable vinyl substituent. This cascade cyclization approach of allylic alcohols involving C-H allylation has not been reported previously. The commercially available allylic alcohol substrates, the only by-product of water, and the used terminal oxidant of O2 provide environmentally benign advantages.

MicroRNA-374b inhibits breast cancer progression through regulating CCND1 and TGFA genes.

Emerging evidence indicates that microRNAs (miRNAs) play a critical role in breast cancer development. We recently reported that a higher expression of miR-374b in tumor tissues was associated with a better disease-free survival of triple-negative breast cancer (TNBC). However, the functional significance and molecular mechanisms underlying the role of miR-374b in breast cancer are largely unknown. In this current study, we evaluated the biological functions and potential mechanisms of miR-374b in both TNBC and non-TNBC. We found that miR-374b was significantly downregulated in breast cancer tissues, compared to adjacent tissues. MiR-374b levels were also lower in breast cancer cell lines, as compared to breast epithelial cells. In vitro and in vivo studies demonstrated that miR-374b modulates the malignant behavior of breast cancer cells, such as cell proliferation in 2D and 3D, cell invasion ability, colony-forming ability and tumor growth in mice. By using bioinformatics tools, we predicted that miR-374b plays a role in breast cancer cells through negatively regulating cyclin D1 (CCND1) and transforming growth factor alpha (TGFA). We further confirmed that CCND1 and TGFA contribute to the malignant behavior of breast cancer cells in vitro and in vivo. Our rescue experiments showed that overexpressing CCND1 or TGFA reverses the phenotypes caused by miR-374b overexpression. Taken together, our studies suggest that miR-374b modulates malignant behavior of breast cancer cells by negatively regulating CCND1 and TGFA genes. The newly identified miR-374b-mediated CCND1 and TGFA gene silencing may facilitate a better understanding of the molecular mechanisms of breast cancer progression.

Mer regulates microglial/macrophage M1/M2 polarization and alleviates neuroinflammation following traumatic brain injury.

BACKGROUND: Traumatic brain injury (TBI) is a leading cause of death and disability worldwide. Microglial/macrophage activation and neuroinflammation are key cellular events following TBI, but the regulatory and functional mechanisms are still not well understood. Myeloid-epithelial-reproductive tyrosine kinase (Mer), a member of the Tyro-Axl-Mer (TAM) family of receptor tyrosine kinases, regulates multiple features of microglial/macrophage physiology. However, its function in regulating the innate immune response and microglial/macrophage M1/M2 polarization in TBI has not been addressed. The present study aimed to evaluate the role of Mer in regulating microglial/macrophage M1/M2 polarization and neuroinflammation following TBI. METHODS: The controlled cortical impact (CCI) mouse model was employed. Mer siRNA was intracerebroventricularly administered, and recombinant protein S (PS) was intravenously applied for intervention. The neurobehavioral assessments, RT-PCR, Western blot, magnetic-activated cell sorting, immunohistochemistry and confocal microscopy analysis, Nissl and Fluoro-Jade B staining, brain water content measurement, and contusion volume assessment were performed. RESULTS: Mer is upregulated and regulates microglial/macrophage M1/M2 polarization and neuroinflammation in the acute stage of TBI. Mechanistically, Mer activates the signal transducer and activator of transcription 1 (STAT1)/suppressor of cytokine signaling 1/3 (SOCS1/3) pathway. Inhibition of Mer markedly decreases microglial/macrophage M2-like polarization while increases M1-like polarization, which exacerbates the secondary brain damage and sensorimotor deficits after TBI. Recombinant PS exerts beneficial effects in TBI mice through Mer activation. CONCLUSIONS: Mer is an important regulator of microglial/macrophage M1/M2 polarization and neuroinflammation, and may be considered as a potential target for therapeutic intervention in TBI.

Ceria nanoparticles ameliorate white matter injury after intracerebral hemorrhage: microglia-astrocyte involvement in remyelination.

BACKGROUND: Intracerebral hemorrhage (ICH) can induce excessive accumulation of reactive oxygen species (ROS) that may subsequently cause severe white matter injury. The process of oligodendrocyte progenitor cell (OPC) differentiation is orchestrated by microglia and astrocytes, and ROS also drives the activation of microglia and astrocytes. In light of the potent ROS scavenging capacity of ceria nanoparticles (CeNP), we aimed to investigate whether treatment with CeNP ameliorates white matter injury by modulating ROS-induced microglial polarization and astrocyte alteration. METHODS: ICH was induced in vivo by collagenase VII injection. Mice were administered with PLX3397 for depleting microglia. Primary microglia and astrocytes were used for in vitro experiments. Transmission electron microscopy analysis and immunostaining were performed to verify the positive effects of CeNP in remyelination and OPC differentiation. Flow cytometry, real-time polymerase chain reaction, immunofluorescence and western blotting were used to detect microglia polarization, astrocyte alteration, and the underlying molecular mechanisms. RESULTS: CeNP treatment strongly inhibited ROS-induced NF-κB p65 translocation in both microglia and astrocytes, and significantly decreased the expression of M1 microglia and A1 astrocyte. Furthermore, we found that CeNP treatment promoted remyelination and OPC differentiation after ICH, and such effects were alleviated after microglial depletion. Interestingly, we also found that the number of mature oligodendrocytes was moderately increased in ICH + CeNP + PLX3397-treated mice compared to the ICH + vehicle + PLX3397 group. Therefore, astrocytes might participate in the pathophysiological process. The subsequent phagocytosis assay indicated that A1 astrocyte highly expressed C3, which could bind with microglia C3aR and hinder microglial engulfment of myelin debris. This result further replenished the feedback mechanism from astrocytes to microglia. CONCLUSION: The present study reveals a new mechanism in white matter injury after ICH: ICH induces M1 microglia and A1 astrocyte through ROS-induced NF-κB p65 translocation that hinders OPC maturation. Subsequently, A1 astrocytes inhibit microglial phagocytosis of myelin debris via an astrocytic C3-microglial C3aR axis. Polyethylene glycol-CeNP treatment inhibits this pathological process and ultimately promotes remyelination. Such findings enlighten us that astrocytes and microglia should be regarded as a functional unit in future works.

Discovery of LAMP-2A as potential biomarkers for glioblastoma development by modulating apoptosis through N-CoR degradation.

BACKGROUND: Lysosome-associated membrane protein type 2A (LAMP-2A) is the key component of chaperone-mediated autophagy (CMA), a cargo-selective lysosomal degradation pathway. Aberrant LAMP-2A expression and CMA activation have been demonstrated in various human malignancies. The study focusing on the intrinsic role of LAMP-2A and CMA in glioblastoma (GBM), and downstream mechanism could provide valuable insight into the pathogenesis and novel therapeutic modality of GBM. METHODS: The levels of LAMP-2A, nuclear receptor co-repressor (N-CoR), unfolded protein response (UPR) and apoptosis were examined in clinical samples. LAMP-2A siRNA and shRNA were constructed to manipulate CMA activation. The role of CMA and downstream mechanism through degradation of N-CoR and arresting UPR mediated apoptosis were explored in GBM cells and nude mouse xenograft model. RESULTS: Elevated LAMP-2A and associated decreased N-CoR expression were observed in GBM as compared with peritumoral region and low-grade glioma. Inhibited UPR and apoptosis were observed in GBM with high LAMP-2A expression. In vitro study demonstrated co-localization and interaction between LAMP-2A and N-CoR. LAMP-2A silencing up-regulated N-CoR and aroused UPR pathway, leading to apoptosis, while N-CoR silencing led to an opposite result. In vivo study further confirmed that LAMP-2A inhibition arrested tumor growth by promoting apoptosis. CONCLUSIONS: Our results demonstrated the central role of CMA in mediating N-CoR degradation and protecting GBM cells against UPR and apoptosis, and provided evidence of LAMP-2A as potential biomarker. Further research focusing on CMA with other tumorigenic process is needed and selective modulators of LAMP-2A remain to be investigated to provide a novel therapeutic strategy for GBM. Video Abstract.

Preoperative controlling nutritional status (CONUT) score is a prognostic factor for early-stage cervical cancer patients with high-risk factors.

OBJECTIVE: The goal of the study was to confirm whether preoperative controlling nutritional status (CONUT) was a prognostic factor in early-stage cervical cancer patients with high-risk factors after surgery and postoperative concurrent chemoradiotherapy (CCRT). METHODS: Between 2004 and 2015, a total of 698 patients who were treated with surgery and postoperative CCRT were included in this retrospective study. The prescribed dose for postoperative radiotherapy was 45-50.4 Gy in 25-28 fractions and the concurrent chemotherapy regimen contained cisplatin or paclitaxel. Based on the receiver operating characteristic (ROC) analysis, the patients were classified into low (<3) and high (≥3) CONUT groups. RESULTS: Of all study patients, 471 (67.5%) patients were included in the low CONUT group. The low CONUT group had significantly better 5-year disease-free survival (DFS) and overall survival (OS) than the high CONUT group (p<0.001 and p = 0.001, respectively). A high CONUT score was significantly associated with lymph node metastasis, parametrial invasion, and poorer nutritional status, including lower body mass index (BMI) and lower prognostic nutritional index (PNI) score (p<0.05, respectively). The CONUT score was an independent predictor of DFS and OS in multivariate analysis. Notably, the CONUT score still efficiently stratified DFS in the high PNI score group (P = 0.001). CONCLUSIONS: The preoperative high CONUT scores indicated poor prognosis for early-stage cervical cancer patients with high-risk factors treated with surgery and postoperative CCRT. In clinical practice, patients with high CONUT score should be considered to receive consolidation chemotherapy.

Deep venous drainage variant rate and degree may be higher in patients with perimesencephalic than in non-perimesencephalic angiogram-negative subarachnoid hemorrhage.

OBJECTIVES: The basal vein of Rosenthal (BVR) variant is a potential origin of bleeding in angiogram-negative subarachnoid hemorrhage (AN-SAH). We compared the rate and degree of BVR variants in patients with perimesencephalic AN-SAH (PAN-SAH) and non-perimesencephalic AN-SAH (NPAN-SAH). METHODS: We retrospectively reviewed the records of AN-SAH patients admitted to our hospital between 2013 and 2018. The associations between variables (baseline characteristics, clinical and radiological data, and outcome) with bleeding patterns and degree of BVR variants were analyzed. Additionally, potential predictors of positive findings on repeated digital-subtracted angiogram (DSA), rebleeding, delayed cerebral infarction (DCI), and poor outcome were further studied. RESULTS: A total of 273 patients with AN-SAH were included. The incidence rate and degree of BVR variants were significantly higher in PAN-SAH patients compared with those in NPAN-SAH patients (p < 0.001). Patients with normal bilateral BVRs are more likely to have a severe prognosis and diffused blood distribution (p < 0.05). We found an increased rate of positive findings on repeated DSA, DCI, rebleeding, and poor outcome at 3 months and 1 year after discharge (all p < 0.05) in patients with bilateral normal BVRs. Bilateral normal BVRs were considered a risk factor (predictor) of positive findings on repeated DSA, rebleeding, and poor outcome (all p < 0.05). CONCLUSIONS: PAN-SAH patients have a higher rate and degree of BVR variants compared with patients with NPAN-SAH. Those AN-SAH patients with bilateral normal BVRs are more likely to be of arterial origin and are at risk of suffering from rebleeding and a poor outcome. KEY POINTS: • Patients with PAN-SAH have a higher rate and degree of BVR variants compared with patients with NPAN-SAH, which suggested that AN-SAH patients with normal BVRs are more likely to originate from arterial bleeding. • AN-SAH patients with normal BVRs are more likely to have positive findings on repeated DSA examinations, as well as an increased incidence of rebleeding and poor outcome, which may assist and guide neurologists in selecting treatment.

Post-operative small pelvic intensity-modulated radiation therapy for early-stage cervical cancer with intermediate-risk factors: efficacy and toxicity.

OBJECTIVE: The aim of the present study was to retrospectively evaluate the toxicity and efficacy of post-operative small pelvic intensity-modulated radiotherapy in early-stage cervical cancer patients with intermediate-risk factors. METHODS: Between 2012 and 2016, 151 patients who had cervical cancer (International Federation of Gynecology and Obstetrics stage I-IIA) with intermediate-risk factors were treated with post-operative small pelvic intensity-modulated radiotherapy. The median dose of 50.4 Gy in 28 fractions with small pelvic intensity-modulated radiotherapy was prescribed to the planning target volume. The intensity-modulated radiotherapy technique used was conventional fixed-field intensity-modulated radiotherapy or helical tomotherapy. RESULTS: The median follow-up was 37 months. The 3-year disease-free survival and overall survival rates were 89 and 96%, respectively. A total of 144 patients (95.3%) were alive at the last follow-up. In total, 6 patients (3.9%) had recurrence: locoregional recurrence in 3 patients (2%), distant metastasis in 2 (1.3%), and both in 1 (0.6%). Diarrhoea was the most common acute toxicity. There were no patients suffering from acute or late grade ≥ 3 toxicity. Only 4 patients (2.6%) had late grade 2 toxicities. CONCLUSIONS: For early-stage cervical cancer patients with intermediate-risk factors, post-operative small pelvic intensity-modulated radiotherapy was safe and well tolerated. The rates of acute and late toxicities were quite satisfactory.

Asymmetric Palladium-Catalyzed C-H Functionalization Cascade for Synthesis of Chiral 3,4-Dihydroisoquinolones.

A palladium-catalyzed C-H functionalization/intramolecular asymmetric allylation cascade of N-sulfonyl benzamides with 1,3-dienes has been developed. In the presence of a chiral pyridine-oxazoline ligand, this protocol enables the synthesis of chiral 3,4-dihydroisoquinolones in yields of up to 83% with enantioselectivities of up to 96%, using environmentally friendly air as the terminal oxidant.

Vinylogous Elimination/Heck Coupling/Allylation Domino Reactions: Access to 2-Substituted 2,3-Dihydrobenzofurans and Indolines.

A highly regio- and stereoselctive palladium-catalyzed domino reaction of functionalized aryl allyl ethers has been developed. Various aryl allyl ethers derived from the phosphine-catalyzed addition of electron-deficient allenes with phenol are found to be efficient substrates for the synthesis of 2-substituted 2,3-dihydrobenzofurans and indolines. It is the first example of aryl allyl ether used as an ideal and practical precursor of hard to get functionalized 1,3-butadiene for the heterocyclic compound synthesis.

[Analysis of TGFBI gene mutation in a Chinese pedigree affected with lattice corneal dystrophy].

OBJECTIVE: To explore the clinical features and mutation of TGFBI gene in a Chinese pedigree affected with lattice corneal dystrophy (LCD). METHODS: Genomic DNA was extracted from 35 members including 11 patients from the pedigree. The 17 exons and splicing region of introns of the TGFBI gene were amplified by PCR. The products were directly sequenced and compared with GenBank database to identify potential mutation. Bioinformatic analysis was carried out to predict the effect of mutation on proteins. RESULTS: A heterozygous mutation (p.R124C) was found in exon 4 of the TGFBI gene in all patients from the pedigree but not among unaffected members. The mode of inheritance of corneal dystrophy in this pedigree was identified as autosomal dominant. Bioinformatics analysis predicted that the p.R124C mutation may be functionally deleterious. The phenotype of corneal dystrophy in the pedigree was determined to be LCD I type. CONCLUSION: The p.R124C mutation of the TGFBI gene probably underlies the pathogenesis of LCD in this Chinese pedigree. Genetic testing can facilitate proper diagnosis of this type of corneal dystrophy.

Neuroprotective Mechanisms of Melatonin in Hemorrhagic Stroke.

Hemorrhagic stroke which consists of subarachnoid hemorrhage and intracerebral hemorrhage is a dominant cause of death and disability worldwide. Although great efforts have been made, the physiological mechanisms of these diseases are not fully understood and effective pharmacological interventions are still lacking. Melatonin (N-acetyl-5-methoxytryptamine), a neurohormone produced by the pineal gland, is a broad-spectrum antioxidant and potent free radical scavenger. More importantly, there is extensive evidence demonstrating that melatonin confers neuroprotective effects in experimental models of hemorrhagic stroke. Multiple molecular mechanisms such as antioxidant, anti-apoptosis, and anti-inflammation, contribute to melatonin-mediated neuroprotection against brain injury after hemorrhagic stroke. This review article aims to summarize current knowledge regarding the beneficial effects of melatonin in experimental models of hemorrhagic stroke and explores the underlying mechanisms. We propose that melatonin is a promising neuroprotective candidate that is worthy of further evaluation for its potential therapeutic applications in hemorrhagic stroke.

Analysis of the VSX1 gene in sporadic keratoconus patients from China.

BACKGROUND: Keratoconus normally presents as a sporadic disease. Although different studies have found sequence variants of the visual system homeobox 1 (VSX1) gene associated with keratoconus in humans, no research has detected such variants in sporadic keratoconus patients from China. To investigate the possibility of VSX1 being a candidate susceptibility gene for Chinese patients with sporadic keratoconus, we performed sequence screening of this gene in such patients. METHODS: Whole DNA was obtained from the leukocytes in the peripheral venous blood of 50 patients with sporadic keratoconus and 50 control subjects without this ocular disorder. Polymerase chain reaction single-strand conformation polymorphism analysis and direct DNA sequencing technology were used to detect sequence variation in the five exons and splicing regions of the introns of the VSX1 gene. The sequencing results were analyzed using DNAstar software. RESULTS: One novel missense heterozygous sequence variant (p.Arg131Pro) was found in the first exon of the VSX1 gene in one keratoconus patient. Another heterozygous sequence variant (p.Gly160Val) in the second exon was found in two keratoconus patients. These variants were not detected in the control subjects. In the third intron of the VSX1 gene, c.8326G > A nucleotide substitution (including heterozygous and homozygous change) was also discovered. The frequency of this variation did not differ significantly between patients and controls, it should belong to single-nucleotide polymorphism of the VSX1 gene. Bioinformatic analysis also predicted that one missense sequence variation (p.Arg131Pro) may not cause a pathogenic change. CONCLUSIONS: In this study, we added one novel missense sequence variation (p.Arg131Pro) in the coding region of the VSX1 gene to the range of VSX1 coding region variations observed in patients with sporadic keratoconus from China. Our work suggests that VSX1 sequence variants might be involved in the pathogenesis of sporadic keratoconus, but their precise role in disease causation requires further investigation.

The Polarization States of Microglia in TBI: A New Paradigm for Pharmacological Intervention.

Traumatic brain injury (TBI) is a serious medical and social problem worldwide. Because of the complex pathophysiological mechanisms of TBI, effective pharmacotherapy is still lacking. The microglial cells are resident tissue macrophages located in the brain and have two major polarization states, M1 phenotype and M2 phenotype, when activated. The M1 phenotype is related to the release of proinflammatory cytokines and secondary brain injury, while the M2 phenotype has been proved to be responsible for the release of anti-inflammation cytokines and for central nervous system (CNS) repair. In animal models, pharmacological strategies inhibiting the M1 phenotype and promoting the M2 phenotype of microglial cells could alleviate cerebral damage and improve neurological function recovery after TBI. In this review, we aimed to summarize the current knowledge about the pathological significance of microglial M1/M2 polarization in the pathophysiology of TBI. In addition, we reviewed several drugs that have provided neuroprotective effects against brain injury following TBI by altering the polarization states of the microglia. We emphasized that future investigation of the regulation mechanisms of microglial M1/M2 polarization in TBI is anticipated, which could contribute to the development of new targets of pharmacological intervention in TBI.

[Analysis of TGFBI gene mutation in a Chinese family affected with Reis-Bucklers corneal dystrophy].

OBJECTIVE: To analyze the clinical features and TGFBI gene mutation in a Chinese family affected with Reis-Bucklers corneal dystrophy. METHODS: Genomic DNA was extracted from 53 members including 9 patients from the family. The 17 exons and splice region of introns of the TGFBI gene were amplified by PCR and directly sequenced. All family members were subjected to ophthalmologic examination. RESULTS: A heterozygous mutation (R124L) was found in exon 4 of the TGFBI gene among all patients from the family. The same mutation was not found among unaffected family members. The inheritance pattern of the family was identified as autosomal dominant, and the Reis-Bucklers corneal dystrophy in the family was diagnosed as the geographic type. CONCLUSION: The R124L mutation of the TGFBI gene probably underlies the pathogenesis of Reis-Bucklers corneal dystrophy in this Chinese family. Molecular genetic approach is useful for the proper diagnosis of this type of corneal dystrophy.

MiR-374a suppresses lung adenocarcinoma cell proliferation and invasion by targeting TGFA gene expression.

Aberrant expression of miR-374a has been reported in several types of human cancers, including lung cancer. However, the functional significance and molecular mechanisms underlying the role of miR-374a in lung cancer remain largely unknown. We found that the expression of miR-374a was significantly downregulated in lung adenocarcinoma tissues compared to adjacent normal lung tissues in samples included in The Cancer Genome Atlas. Functional studies revealed that overexpression of miR-374a led to inhibition of lung adenocarcinoma cell proliferation, migration and invasion and that miR-374a negatively regulated transforming growth factor-alpha (TGFA) gene expression by directly targeting the 3'-UTR of TGFA mRNA. Treating lung adenocarcinoma cells with TGF-α neutralizing antibody resulted in suppression of cell proliferation and invasion, which mimicked the action of miR-374a. Additionally, TGFA gene expression was significantly higher in tumor tissues compared to adjacent normal tissue and high TGFA gene expression strongly correlated with poor survival in patients with lung adenocarcinoma. Taken together, our studies suggest that miR-374a suppresses lung adenocarcinoma cell proliferation and invasion via targeting TGFA gene expression. Our findings may provide novel treatment strategies for lung adenocarcinoma patients.

The autophagy-lysosomal system in subarachnoid haemorrhage.

The autophagy-lysosomal pathway is a self-catabolic process by which dysfunctional or unnecessary intracellular components are degraded by lysosomal enzymes. Proper function of this pathway is critical for maintaining cell homeostasis and survival. Subarachnoid haemorrhage (SAH) is one of the most devastating forms of stroke. Multiple pathogenic mechanisms, such as inflammation, apoptosis, and oxidative stress, are all responsible for brain injury and poor outcome after SAH. Most recently, accumulating evidence has demonstrated that the autophagy-lysosomal pathway plays a crucial role in the pathophysiological process after SAH. Appropriate activity of autophagy-lysosomal pathway acts as a pro-survival mechanism in SAH, while excessive self-digestion results in cell death after SAH. Consequently, in this review article, we will give an overview of the pathophysiological roles of autophagy-lysosomal pathway in the pathogenesis of SAH. And approaching the molecular mechanisms underlying this pathway in SAH pathology is anticipated, which may ultimately allow development of effective therapeutic strategies for SAH patients through regulating the autophagy-lysosomal machinery.

Melatonin attenuates neuronal apoptosis through up-regulation of K(+) -Cl(-) cotransporter KCC2 expression following traumatic brain injury in rats.

Traumatic brain injury (TBI) initiates a complex cascade of neurochemical and signaling changes that leads to neuronal apoptosis, which contributes to poor outcomes for patients with TBI. The neuron-specific K(+) -Cl(-) cotransporter-2 (KCC2), the principal Cl(-) extruder in adult neurons, plays an important role in Cl(-) homeostasis and neuronal function. This present study was designed to investigate the expression pattern of KCC2 following TBI and to evaluate whether or not melatonin is able to prevent neuronal apoptosis by modulating KCC2 expression in a Sprague Dawley rat controlled cortical impact model of TBI. The time course study showed decreased mRNA and protein expression of KCC2 in the ipsilateral peri-core parietal cortex after TBI. Double immunofluorescence staining demonstrated that KCC2 is located in the plasma membrane of neurons. In addition, melatonin (10 mg/kg) was injected intraperitoneally at 5 minutes and repeated at 1, 2, 3, and 4 hours after brain trauma, and brain samples were extracted 24 hours after TBI. Compared to the vehicle group, melatonin treatment altered the down-regulation of KCC2 expression in both mRNA and protein levels after TBI. Also, melatonin treatment increased the protein levels of brain-derived neurotrophic factor (BDNF) and phosphorylated extracellular signal-regulated kinase (p-ERK). Simultaneously, melatonin administration ameliorated cortical neuronal apoptosis, reduced brain edema, and attenuated neurological deficits after TBI. In conclusion, our findings suggested that melatonin restores KCC2 expression, inhibits neuronal apoptosis and attenuates secondary brain injury after TBI, partially through activation of BDNF/ERK pathway.