Extracellular silica nanocoat formed by layer-by-layer (LBL) self-assembly confers aluminum resistance in root border cells of pea (Pisum sativum).

BACKGROUND: Soil acidity (and associated Al toxicity) is a major factor limiting crop production worldwide and threatening global food security. Electrostatic layer-by-layer (LBL) self-assembly provides a convenient and versatile method to form an extracellular silica nanocoat, which possess the ability to protect cell from the damage of physical stress or toxic substances. In this work, we have tested a hypothesis that extracellular silica nanocoat formed by LBL self-assembly will protect root border cells (RBCs) and enhance their resistance to Al toxicity. RESULTS: Scanning electron microscopy (SEM) and X-ray photoelectron spectroscopy (XPS) were used to compare the properties of RBCs surface coated with nanoshells with those that were exposed to Al without coating. The accumulation of Al, reactive oxygen species (ROS) levels, and the activity of mitochondria were detected by a laser-scanning confocal microscopy. We found that a crystal-like layer of silica nanoparticles on the surface of RBCs functions as an extracellular Al-proof coat by immobilizing Al in the apoplast and preventing its accumulation in the cytosol. The silica nanoshells on the RBCs had a positive impact on maintaining the integrity of the plasma and mitochondrial membranes, preventing ROS burst and ensuring higher mitochondria activity and cell viability under Al toxicity. CONCLUSIONS: The study provides evidence that silica nanoshells confers RBCs Al resistance by restraining of Al in the silica-coat, suggesting that this method can be used an efficient tool to prevent multibillion-dollar losses caused by Al toxicity to agricultural crop production.

Somatostatin Reduces the Acute Lung Injury of Mice via Increasing the Affinity of Glucocorticoid Receptor.

BACKGROUND/AIMS: Although it has been reported that somatostatin (SOM) upregulated the level of 90-kD heat shock protein (Hsp90), which participates in the inflammatory regulation by its client proteins, such as glucocorticoid receptor (GR), it remains unclear if it has a protective role against acute lung injury (ALI). METHODS: ALI model was established by the injection of oleic acid (OA) into the tail vein of mice. Lung injury was assessed by histological analysis, lung water content and arterial blood gases. The levels of Hsp90 and GR, the binding capacity and the affinity of GR were examined. RESULTS: It was showed that pretreatment with SOM significantly increased Hsp90 levels and alleviated lung injuries in OA-injected mice. Furthermore, SOM increased the GR expression and improved the affinity of the GR in animals with lung injury. However, little alteration was found in the maximum binding capacity of the GR in mice with or without SOM. CONCLUSION: The data indicate SOM exerts a protective effect by increasing Hsp90 abundant and further enhancing the affinity of the GR. The beneficial effects of SOM treatment provide a new strategy for modulation of GR efficiency and alleviation of acute lung injury.

Chronic caffeine exposure attenuates blast-induced memory deficit in mice.

OBJECTIVE: To investigate the effects of three different ways of chronic caffeine administration on blast- induced memory dysfunction and to explore the underlying mechanisms. METHODS: Adult male C57BL/6 mice were used and randomly divided into five groups: control: without blast exposure, con-water: administrated with water continuously before and after blast-induced traumatic brain injury (bTBI), con-caffeine: administrated with caffeine continuously for 1 month before and after bTBI, pre-caffeine: chronically administrated with caffeine for 1 month before bTBI and withdrawal after bTBI, post-caffeine: chronically administrated with caffeine after bTBI. After being subjected to moderate intensity of blast injury, mice were recorded for learning and memory performance using Morris water maze (MWM) paradigms at 1, 4, and 8 weeks post-blast injury. Neurological deficit scoring, glutamate concentration, proinflammatory cytokines production, and neuropathological changes at 24 h, 1, 4, and 8 weeks post-bTBI were examined to evaluate the brain injury in early and prolonged stages. Adenosine A1 receptor expression was detected using qPCR. RESULTS: All of the three ways of chronic caffeine exposure ameliorated blast-induced memory deficit, which is correlated with the neuroprotective effects against excitotoxicity, inflammation, astrogliosis and neuronal loss at different stages of injury. Continuous caffeine treatment played positive roles in both early and prolonged stages of bTBI; pre-bTBI and post-bTBI treatment of caffeine tended to exert neuroprotective effects at early and prolonged stages of bTBI respectively. Up-regulation of adenosine A1 receptor expression might contribute to the favorable effects of chronic caffeine consumption. CONCLUSION: Since caffeinated beverages are widely consumed in both civilian and military personnel and are convenient to get, the results may provide a promising prophylactic strategy for blast-induced neurotrauma and the consequent cognitive impairment.

The glucocorticoid dexamethasone inhibits U937 cell adhesion and neutrophil release via RhoA/ROCK1-dependent and independent pathways.

AIMS: The aim of the present study was to investigate the role of the Ras homolog family member A (RhoA)/Rho-associated coiled-coil-containing protein kinase 1 (ROCK1) signaling pathway in the inhibition of inflammatory responses by the glucocorticoid dexamethasone (Dex). METHODS: The inhibitory effects of Dex and Rho-kinase inhibitor fasudil (Fas) on phorbol ester-induced release of O2(-) and MPO from neutrophils and on U937 mononuclear cell adhesion were examined along with the expression and activity levels of RhoA and ROCK1. RESULTS: High doses of Dex rapidly inhibited the release of O2(-) and myeloperoxidase (MPO) from neutrophils and the adhesion of U937 cells, while Fas was only found to inhibit U937 cell adhesion. Additionally, Dex suppressed ROCK1 activity. However, Dex had no effects on ROCK1 or RhoA expression levels or on RhoA activity. Neither the glucocorticoid receptor antagonist mifepristone (RU-486) nor the protein synthesis inhibitor cycloheximide (CHX) was able to suppress the effects of Dex (p>0.05). CONCLUSIONS: The present findings indicate that Dex suppressed neutrophil release through ROCK1-independent mechanisms and inhibited the adhesion of U937 mononuclear cells through ROCK1-dependent non-genomic mechanisms that did not involve RhoA.

Clinical outcomes of second-line chemotherapy in patients with advanced pancreatic adenocarcinoma: a real-world study.

OBJECTIVE: Little progress has been made in recent years using first-line chemotherapy, including gemcitabine combined with nab-paclitaxel, FOLFIRINOX, and NALIRIFOX, for advanced pancreatic adenocarcinoma (APC). In addition, the optimal second-line chemotherapy regimen has not been determined. This study aimed to compare the effectiveness of different types of second-line chemotherapy for APC. METHODS: Patients with APC who received first-line treatment from January 2008 to January 2021 were considered eligible for this retrospective analysis. The primary and secondary endpoints were overall survival (OS) and progression-free survival (PFS), respectively. RESULTS: Four hundred and thirty-seven and 617 patients were treated with 5-fluorouracil- and gemcitabine-based chemotherapy as first-line treatment, respectively. Demographic and clinical features, except age and liver metastasis, were comparable between the two groups (P < 0.05). The median OS was 8.8 and 7.8 months in patients who received a 5-fluorouracil- and gemcitabine-based combined regimen for first-line therapy, respectively (HR = 1.244, 95% CI = 1.090-1.419; P < 0.001). The median OS was 5.6 and 1.9 months in patients who received second-line chemotherapy and supportive care, respectively (HR = 0.766, 95% CI = 0.677-0.867; P < 0.001). The median PFS was not significantly differently between gemcitabine or 5-fluorouracil monotherapy and combination therapy. CONCLUSIONS: A 5-fluorouracil- or gemcitabine-based combined regimen was shown to be as effective as a single 5-fluorouracil or gemcitabine regimen as second-line therapy for patients with APC.

The biomineralization of silica induced stress tolerance in plants: a case study for aluminum toxicity.

Biomineralization in plant roots refers to the process of cell-induced self-assembly to form nanostructures on the root surface. Silicon (Si) is the second most abundant element in soils, and beneficial to plant growth. Meanwhile, silicon is shown to participate in the process of biomineralization, which is useful for improving mechanical strength and alleviating biotic and abiotic stress, for example silicic acid polymerizes to form amorphous silica (SiO2-nH2O) in the process of growing to resist fungi and environmental stress. This process alters physical and chemical properties of cell wall. However, the mechanistic basis of this process remains unclear. Aluminum toxicity is a major constraint affecting plant performance in acid soil. This paper summarizes recent research advances in the field of plant biomineralization and describes the effects of silicon biomineralization on plant aluminum tolerance and its adaptive significance, using aluminum toxicity as a case study.

Ski, a modulator of wound healing and scar formation in the rat skin and rabbit ear.

We recently demonstrated that Ski is a novel wound healing-related factor that promotes fibroblast proliferation and inhibits collagen secretion. Here, we show that increasing local Ski expression by gene transfer not only significantly accelerated wound healing by relieving inflammation, accelerating re-epithelialization and increasing formation of granulation tissue, but also reduced scar formation by decreasing collagen production in rat dermal wounds. Similarly, ski gene transfer accelerated wound healing, reduced the protuberant height and volume of scars and increased collagen maturity in a hypertrophic scar model in the rabbit ear. Conversely, reducing Ski expression in the wound by RNA interference resulted in significantly slower wound healing and increased scar area in rat dermal wounds. We demonstrated that these effects of Ski are associated with transforming growth factor-ß-mediated signalling pathways through both Smad2/3-dependent and Smad-independent pathways. Together, our results define a dual role for Ski in promoting wound healing and alleviating scar formation, identifying a new target for therapeutic approaches to preventing scar hyperplasia and accelerating wound healing.

Roles of Rho-associated coiled-coil protein kinase in multiple cell behaviors.

Rho-associated coiled-coil protein kinase (ROCK) is a serine/threonine kinase that belongs to AGC family of kinases. By inducing the formation of stress fibers and reorganizing the cytoskeleton, it is involved in many biological behaviors of cells including cell contraction, cell migration, cell division, and morphological changes, and thus exerts important roles in regulating the multiple functions of cells.

Human umbilical cord mesenchymal stem cells-derived exosomes for treating traumatic pancreatitis in rats.

BACKGROUND: The therapeutic and protective effects of human umbilical cord mesenchymal stem cells-exosomes (hucMSC-Exs) on traumatic pancreatitis (TP) remain unknown. Here, we established a rat model of TP and evaluated and compared the therapeutic effects of hUC-MSCs and hucMSC-Exs. METHODS: HucMSC-Exs were obtained by ultracentrifugation and identified using transmission electron microscopy and western blot analysis. TP rats were treated by tail vein injection of hUC-MSCs and hucMSC-Exs. Their homing in rats was observed by performing fluorescence microscopy. The degree of pancreatic tissue damage was assessed by HE staining, the expression levels of amylase, lipase, and inflammatory cytokines were detected by ELISA, apoptosis was detected by TUNEL assay, and the expression levels of various apoptosis-related proteins were detected by western-blot. The expression levels of apoptosis-related molecular markers were detected by RT-qPCR. RESULTS: The colonization of exosomes was observed in pancreatic tissue. Compared to TP group, the histopathological score of pancreas was significantly decreased in the TP + hUC-MSCs group and TP + hucMSC-Exs group (P < 0.05). Compared to TP group, the activity of serum amylase and lipase was significantly decreased (P < 0.05). The expression levels of IL-6 and TNF-α were significantly decreased, while those of IL-10 and TGF-ß were significantly increased (P < 0.05). The apoptosis index of the TP group was significantly increased (P < 0.05), whereas that of the TP + hUC-MSCs and TP + hucMSC-Exs groups was significantly decreased (P < 0.05). Compared to TP group, the expression levels of Bax, Bcl-2, and Caspase-3 were significantly decreased in the TP + hUC-MSCs group and TP + hucMSC-Exs group (P < 0.05). CONCLUSION: HucMSC-Exs can colonize injured pancreatic tissue, inhibit the apoptosis of acinar cells, and control the systemic inflammatory response to facilitate the repair of pancreatic tissue.

Genome-wide identification and expression analyses of late embryogenesis abundant (LEA) gene family in tobacco (Nicotiana tabacum L.) reveal their function in abiotic stress responses.

Late embryogenesis abundant (LEA) proteins play an important role in plant growth and response to abiotic stresses. However the late embryogenesis abundant (LEA) gene family in Nicotiana tabacum has not been systematically studied. In this study, 123 NtLEA genes were identified in Nicotiana tabacum, and divided into 8 groups, including LEA_1, LEA_2, LEA_3, LEA_4, LEA_5, LEA_6, DHN (dehydratin) and SMP (Seed Maturation Protein). The LEA_2 group is the most abundant of the NtLEA family. The gene structure, conserved motifs, subcellular localization and physicochemical properties of the NtLEA genes were analyzed. RNA-seq and qPCR analyses showed that the NtLEA genes were significantly induced under two different abiotic stresses and showed different expression patterns. The expression patterns of 35 NtLEA genes responding to ABA and 3 NtLEA genes responding to NaCl abiotic stress, respectively, were characterized. The protein-protein interaction network revealed that most NtLEA proteins (>78%) had the potential function to enhance tobacco resistance to abiotic stress. The transcriptional regulatory network showed that 21 transcription factor families were involved in regulating the expression of the NtLEA genes. These results are beneficial for future studies of the function of the NtLEA genes.

The Function of BBX Gene Family under Multiple Stresses in Nicotiana tabacum.

B-box (BBX) is a zinc finger transcription factor, which is involved in regulating the growth and development of plants and resisting various stresses. In this study, 43 NtBBX genes were identified and divided into five subgroups in tobacco. The members in each subgroup had similar characteristics. The promoter region of NtBBX genes had cis-acting elements related to light response, hormone regulation and stress response. Transcriptome analysis showed that NtBBX30 was significantly up-regulated, and NtBBX12, NtBBX13, NtBBX16 and NtBBX17 were significantly down-regulated under abiotic stresses. The NtBBX genes also responded to the infection of Ralstonia solanacearum. NtBBX9, NtBBX1, NtBBX15 and NtBBX17 showed the greatest response under stresses. The NtBBX genes are expressed in various degrees under different tissues. This research will provide a solid foundation for further study of the biological function of NtBBX genes in tobacco.

Effects of Cadmium Stress on Root and Root Border Cells of Some Vegetable Species with Different Types of Root Meristem.

Cadmium is one of the most toxic heavy metals and can be easily absorbed by plants, affecting root growth. Root border cells (RBCs), that are located in the periphery of the root cap and originate from the root cap meristem, represent a convenient tool to study the toxic effects of Cd on root performance. In this work, vegetables with contrasting types of root apical meristem (RAM) organizations were used. The open RAM organizations included pea and cucumber, and the closed RAM organizations included tomato, chili, and eggplant. The number of RBCs were significantly higher in the species possessing open RAM organization: pea (11,330 cells per root) > cucumber (8200) > tomato (2480) > eggplant (1830) > chili (1320). The same trend was observed for cell viability: pea (61%) > cucumber (59%) > tomato (49%) > eggplant (44%) > chili (42%). Pea and cucumber had higher relative radicle elongation rates and a lower increase in stress-induced accumulation of malondialdehyde (MDA), making them more resistant to Cd stress than the vegetables with close RAM organization. Under Cd treatment, the number and viability of RBCs in vegetables with both types of RAM organization were significantly decreased. However, the decreasing ratio of the number and viability of RBCs in pea and cucumber was higher than in tomato, chili, and eggplant. Taken together, the plants with the open-type RAM are more tolerant to Cd, and it can be speculated that the cadmium tolerance of the vegetables may be correlated with the number and viability of RBCs in response to cadmium stress.

Local glutamate level dictates adenosine A2A receptor regulation of neuroinflammation and traumatic brain injury.

During brain injury, extracellular adenosine and glutamate levels increase rapidly and dramatically. We hypothesized that local glutamate levels in the brain dictates the adenosine-adenosine A(2A) receptor (A(2A)R) effects on neuroinflammation and brain damage outcome. Here, we showed that, in the presence of low concentrations of glutamate, the A(2A)R agonist 3-[4-[2-[[6-amino-9-[(2R,3R,4S,5S)-5-(ethylcarbamoyl)-3,4-dihydroxy-oxolan-2-yl]purin-2-yl]amino]ethyl]phenyl]propanoic acid (CGS21680) inhibited lipopolysaccharide (LPS)-induced nitric oxide synthase (NOS) activity of cultured microglial cells, an effect that was dependent on the protein kinase A (PKA) pathway. However, in high concentrations of glutamate, CGS21680 increased LPS-induced NOS activity in a protein kinase C (PKC)-dependent manner. Thus, increasing the local level of glutamate redirects A(2A)R signaling from the PKA to the PKC pathway, resulting in a switch in A(2A)R effects from antiinflammatory to proinflammatory. In a cortical impact model of traumatic brain injury (TBI) in mice, brain water contents, behavioral deficits, and expression of tumor necrosis factor-alpha, interleukin-1 mRNAs, and inducible NOS were attenuated by administering CGS21680 at post-TBI time when brain glutamate levels were low, or by administering the A(2A)R antagonist ZM241385 [4-(2-{[5-amino-2-(2-furyl)[1,2,4]triazolo[1,5-a][1,3,5]triazin-7-yl]amino}ethyl)phenol] at post-TBI time when brain glutamate levels were elevated. Furthermore, pre-TBI treatment with the glutamate release inhibitor (S)-4C3HPG [(S)-4-carboxy-3-hydroxyphenylglycine] converted the debilitating effect of CGS21680 administered at post-TBI time with high glutamate level to a neuroprotective effect. This further indicates that the switch in the effect of A(2A)R activation in intact animals from antiinflammatory to proinflammatory is dependent on glutamate concentration. These findings identify a novel role for glutamate in modulation of neuroinflammation and brain injury via the adenosine-A(2A)R system.

Monosialotetrahexosylganglioside in the treatment of chronic oxaliplatin-induced peripheral neurotoxicity: TJMUCH-GI-001, a randomised controlled trial.

BACKGROUND: Chronic oxaliplatin-induced peripheral neurotoxicity (OIPN) is the most troublesome and dose-limiting side effect of oxaliplatin. There is no effective treatment for chronic OIPN. We conducted a randomised controlled trial to investigate the efficacy of monosialotetrahexosylganglioside (GM1) in treating chronic OIPN. METHODS: In this single-centre, double-blind, phase Ⅲ trial, gastrointestinal cancer patients with persistent chronic OIPN were randomised in 1:1 ratio to receive either GM1 or placebo at Tianjin Medical University Cancer Institute and Hospital, China. GM1 was dosed at 60 mg daily for every 3 weeks or 40 mg daily for every 2 weeks. Seven- and fourteen- day infusions were administered to concurrent oxaliplatin users and oxaliplatin discontinuation patients, respectively. The primary endpoint was the relief of neurotoxicity (≥30% improvement), measured by a newly developed patient reported outcome measure (MCIPN) based on prior questionnaires including the European Organization for Research and Treatment of Cancer Quality of Life Chemotherapy Induced Peripheral Neuropathy Questionnaire twenty-item scale. Visual analogue score (VAS) was used as another instrument for patients to evaluate the total Chronic OIPN treatment effect. VAS responders (≥30% improvement), double responders (≥30% improvement in both MCIPN and VAS), and high responders (≥50% improvement in the MCIPN total score) were also calculated. The secondary endpoints were safety and quality of life. The additional endpoints are progression-free survival (PFS), disease-free survival (DFS), overall survival (OS), and tumour response. (Trial registration number: NCT02486198 at ClinicalTrials.gov). FINDINGS: Between May 2015 to December 2017, 145 patients were randomly assigned to receive either GM1 (n=73) and placebo (n=72). Majority of the patients in both arms (90% in GM1 and 83% in placebo) continued receiving oxaliplatin on the trial. More patients responded in the GM1 group than in the placebo group (MCIPN responders: 53% vs 14%, VAS responders: 49% vs 22%, double responders: 41% vs 7%, and high responders: 32% vs 13%, all P < ·01). Analyses were also performed in concurrent oxaliplatin users. The results were consistent with those of the whole group. No deleterious effects of GM1 on survival or tumour response were found. There were no ≥G3 GM1-related adverse events. INTERPRETATION: In patients with chronic OIPN, the use of GM1 reduces the severity of chronic OIPN compared with placebo. FUNDING: This work was supported by clinical trial development fund of Tianjin Medical University Cancer Institute and Hospital (No.C1706).

Adenosine A2A receptors in both bone marrow cells and non-bone marrow cells contribute to traumatic brain injury.

Adenosine A2A receptors (A(2A)Rs) in bone marrow-derived cells (BMDCs) are involved in regulation of inflammation and outcome in several CNS injuries; however their relative contribution to traumatic brain injury (TBI) is unknown. In this study, we created a mouse cortical impact model, and BMDC A(2A)Rs were selectively inactivated in wild-type (WT) mice or reconstituted in global A(2A)R knockout (KO) mice (i.e. inactivation of non-BMDC A(2A)Rs) by bone marrow transplantation. When compared with WT mice, selective inactivation of BMDC A(2A)Rs significantly attenuated the neurological deficits, brain water content and cell apoptosis at 24 h post-TBI as global A(2A)R KO did. However, compared with the A(2A)R KO mice, selective reconstitution of BMDC A(2A)Rs failed to reinstate brain injury, indicating the contribution of the non-BMDC A(2A)R to TBI. Furthermore, the protective outcome by selective inactivation of BMDC A(2A)R or broad inactivation of non-BMDC A(2A)Rs was accompanied with reduced CSF glutamate level and suppression of the inflammatory cytokines interleukin-1, or interleukin-1 and tumor necrosis factor-alpha. These findings demonstrate that inactivation of A(2A)Rs in either BMDCs or non-BMDCs is sufficient to confer the protective effect as global A(2A)R KO against TBI, indicating the A(2A)R involvement in TBI by multiple cellular mechanisms of A(2A)R involvement including inhibition of glutamate release and inflammatory cytokine expressions.

Genetic variations of heat shock protein 84 in mice mediate cellular glucocorticoid response.

Heat shock protein 90 (Hsp90), encoded by hsp84 and hsp86 in mice, has been confirmed to modulate glucocorticoid receptor (GR) function; however, the contribution of Hsp90 in glucocorticoid (GC) sensibility/resistance has received less attention. Previously, we found that genetic variations of Hsp84 are related to differences in the in vivo GC-GR responses between BALB/c and C57BL/6 mice suffering from traumatic injury. To evaluate the modulation of Hsp84 polymorphisms on the GC response, we used a cellular heat-stress injury (HSI) model combined with a transgene-plasmid infection approach and assessed HSI-induced cellular damage and GR nuclear translocation, with or without dexamethasone pretreatment. We demonstrated that after HSI, fibroblasts from the C57BL/6 line exhibit higher cellular survival, higher nuclear GR levels and lower lactate dehydrogenase activity compared to those from the BALB/c line. We showed that dexamethasone-rescued HSI-induced damage is accompanied by increasing nuclear GR levels in both lines. Importantly, this protection against HSI was greater in C57BL/6 fibroblasts and was resistant to geldanamycin, a selective inhibitor of Hsp90. Importantly, transfection of the hsp84-transgene from C57BL/6 mice increased the nuclear GR levels and lessened HSI-induced damage in BALB/c fibroblasts. Our data thereby demonstrate that Hsp84 from C57BL/6 mice modulates higher cellular GC-GR responsiveness.

Application of an oligonucleotide microarray-based nano-amplification technique for the detection of fungal pathogens.

BACKGROUND: The traditional techniques for diagnosis of invasive fungal infections in the clinical microbiology laboratory need improvement. These techniques are prone to delay results due to their time-consuming process, or result in misidentification of the fungus due to low sensitivity or low specificity. The aim of this study was to develop a method for the rapid detection and identification of fungal pathogens. METHODS: The internal transcribed spacer two fragments of fungal ribosomal DNA were amplified using a polymerase chain reaction for all samples. Next, the products were hybridized with the probes immobilized on the surface of a microarray. These species-specific probes were designed to detect nine different clinical pathogenic fungi including Candida albicans, Candida tropocalis, Candida glabrata, Candida parapsilosis, Candida krusei, Candida lusitaniae, Candida guilliermondii, Candida keyfr, and Cryptococcus neoformans. The hybridizing signals were enhanced with gold nanoparticles and silver deposition, and detected using a flatbed scanner or visually. RESULTS: Fifty-nine strains of fungal pathogens, including standard and clinically isolated strains, were correctly identified by this method. The sensitivity of the assay for Candida albicans was 10 cells/mL. Ten cultures from clinical specimens and 12 clinical samples spiked with fungi were also identified correctly. CONCLUSIONS: This technique offers a reliable alternative to conventional methods for the detection and identification of fungal pathogens. It has higher efficiency, specificity and sensitivity compared with other methods commonly used in the clinical laboratory.

[Effect of (S)-4C3HPG on brain damage in the acute stage of moderate traumatic brain injury model of mice and underlying mechanism].

The aim of this study is to investigate the effect of (S)-4-carboxy-3-hydroxy-phenylglycine [(S)-4C3HPG], a mixed group I glutamate metabotropic receptor antagonist and a group II agonist, on impairment in a cortical impact model of traumatic brain injury (TBI) in mice and to elucidate the possible mechanisms. Mice were injected (i.p.) with saline, 1 mg/kg (S)-4C3HPG, 5 mg/kg (S)-4C3HPG and 10 mg/kg (S)-4C3HPG (n=10 per group), respectively, at 30 min before moderate TBI. Neurological deficit scores, water content in injured brain and glutamate concentration in cerebral spinal fluid (CSF) were detected at 24 h after TBI. The expressions of tumor necrosis factor-α (TNF-α) and interleukin-1ß (IL-1ß) mRNA in injured cortex were also detected by real-time RT-PCR. The results showed that the neurological deficits and cerebral edema were significantly attenuated in mice pretreated with (S)-4C3HPG (5 and 10 mg/kg respectively) compared with those in mice pretreated with saline. Furthermore, (S)-4C3HPG treatment also decreased the glutamate concentration in CSF and the expressions of TNF-α and IL-1ß mRNA remarkably in a dose-dependent manner. These results suggest that (S)-4C3HPG treatment attenuates cortical impact-induced brain injury possibly via suppression of glutamate release and inhibition of excessive inflammatory cytokine production. These findings highlight the potential benefit of glutamate metabotropic receptor ligand for preventing TBI.

Identification and tissue distribution of a novel rat glucocorticoid receptor splice variant.

Glucocorticoid receptor (GR) is a steroid hormone receptor that has been shown to play important roles in diverse cellular and physiological processes. More and more evidence has revealed that the effects of glucocorticoids are mediated by the glucocorticoid receptor through genomic or nongenomic mechanisms. A growing number of glucocorticoid receptor splice variants have been identified in human tissues, but few are known in rat tissues. In this work, a novel rGR cDNA, called rGRbeta, was cloned from Sprague Dawlay (SD) rat liver. Sequence analysis revealed that the rGRbeta mRNA was 39 base pairs (bp) shorter than the rGR mRNA reported earlier. The deleted segment is located in exon 1 and encodes 13 repeated glutamine residues. Both the rGR and rGRbeta mRNAs were quantitated by Northern blot hybridization using non-homologous glucocorticoid cDNA probes. Results showed that the rGR and rGRbeta mRNAs were most abundant in the lung, the least abundant in the heart, and there were more rGR and rGRbeta mRNAs in the kidney than in the liver. The identification of rGRbeta may contribute to the understanding of the genomic or nongenomic effects of glucocorticoids.

The essential role for c-Ski in mediating TGF-beta1-induced bi-directional effects on skin fibroblast proliferation through a feedback loop.

The bi-directional regulation of TGF-beta1 (transforming growth factor-beta1) on fibroblast proliferation with stimulation at low concentration, but inhibition at high concentration, has important significance during tissue repair. The mechanism has not been defined. c-Ski is a major co-repressor of TGF-beta1/Smad3 signalling; however, the exact role of c-Ski in the bi-directional regulation of fibroblast proliferation remains to be determined. In the present study, we established a dose-effect relationship of bi-directional regulation of TGF-beta1-mediated proliferation in rat skin fibroblasts, and found that c-Ski overexpression promoted fibroblast proliferation by inhibiting Smad3 activity. Importantly, c-Ski expression was decreased at the high concentration of TGF-beta1, but increased at the low concentration of TGF-beta1. This dose-dependent change in TGF-beta1 action did not affect Smad3 phosphorylation or nuclear translocation, but altered Smad3 DNA-binding activity, transcriptional activity and expression of the downstream gene p21 that both increased at the high concentration and decreased at the low concentration. Furthermore, c-Ski overexpression exerted synergistic stimulation with TGF-beta1 at the low concentration, but reversed the inhibitory effect of TGF-beta1 at high concentrations, while knockdown of c-Ski by RNA interference abrogated bi-directional role of TGF-beta1 on fibroblast proliferation. Thus our data reveal a new mechanism for this bi-directional regulation, i.e. c-Ski expression change induced by low or high TGF-beta1 concentration in turn determines the promoting or inhibiting effects of TGF-beta1 on fibroblast proliferation, and suggests an important role of c-Ski that modulates the local availability of TGF-beta1 within the wound repair microenvironment.