To cut or not to cut? A prospective randomized controlled trial on short-term outcomes of the uncut Roux-en-Y reconstruction for gastric cancer.

BACKGROUND: Roux-en-Y (R-Y) anastomoses have been widely used in distal gastrectomy, while the incidence of Roux stasis syndrome remains common. Uncut R-Y anastomosis maintains the neuromuscular continuity, thus avoiding the ectopic pacemaker of the Roux limb and reducing the occurrence of Roux stasis. However, retrospective studies of Uncut R-Y anastomosis remain scarce and randomized controlled trials have not been reported. METHODS: We conducted a randomized controlled trial to compare the surgical safety, nutritional status, and postoperative quality of life (QOL) between uncut and classic Roux-en-Y (R-Y) reconstruction patients. Patients with Stage I gastric cancer were randomly enrolled and underwent laparoscopic distal gastrectomy followed by uncut or classic R-Y reconstruction. Body mass index and blood test were used to evaluate the nutritional status. QOL was evaluated using European Organization for Research and Treatment of Cancer QOL Questionnaire (STO22) and laboratory examinations at postoperative month (POM) 3, 6, 9, and 12. Computed tomography scanning was used to evaluate the skeletal muscle index (SMI) at POM 6 and 12. Endoscopy was performed at POM 12. RESULTS: Operation time, blood loss, time to recovery, complication morbidities, and overall survival were similar between the two groups. Compared with the classic R-Y group, the uncut R-Y group displayed a significantly decreased QOL at POM 9, possibly due to loop recanalization, determined to be occupied 34.2% of the uncut R-Y group. Post-exclusion of recanalization, the QOL was still higher in the classic R-Y group than in the uncut R-Y group, despite their hemoglobin and total protein levels being better than those in the classic R-Y group. Preoperative pre-albumin level and impaired fasting glycemia significantly correlated with the postoperative recanalization. CONCLUSION: We found no significant benefit of uncut over classic R-Y reconstruction which challenges the superiority of the uncut R-Y reconstruction. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT02644148.

Comparison of long-term oncologic outcomes laparoscopy-assisted gastrectomy and open gastrectomy for gastric cancer.

PURPOSE: Laparoscopy-assisted gastrectomy (LAG) is proven by considerable studies as a safe procedure for early gastric cancer (EGC), but its long-term oncologic outcomes in advanced gastric cancer (AGC) have not been well-described. This study aimed at verifying the non-inferiority of LAG in the treatment of EGC and comparing the oncological feasibility of LAG and open gastrectomy (OG) for AGC. METHODS: A total of 209 consecutive patients who underwent LAG or OG with D2 lymph node dissection between December 2008 and November 2012 were included. The survival rate was estimated with the Kaplan-Meier method and the risk factors affecting the survival and recurrence were evaluated with Cox regression models. Subgroup analysis was performed in AGC patients receiving both distal and total gastrectomy. RESULTS: Of 209 patients, 194 (92.8%; mean age, 62.7 years; 56 [28.9%] women) eligible patients were finally enrolled in this study. No significant differences in the number of lymph nodes retrieved and postoperative complications were observed between patients receiving LAG and OG. During a mean follow-up of 58.3 ± 38.1 months (range 0-121 months), the 5-year overall survival and disease-free survival rates were 56.1% and 53.0% for LAG, and 57.7% and 50.9% for OG. In the subgroup analysis for AGC, laparoscopy-assisted distal gastrectomy and total gastrectomy did not result in inferior long-term outcomes, and recurrence was found in 49 patients (31.2%). Age more than 65 years and the advanced tumor stage were independent risk factors of survival. CONCLUSION: LAG is a feasible and safe treatment for gastric cancer, with good oncologic results.

Uncut Roux-en-Y Reconstruction in a Laparoscopic Distal Gastrectomy: A Single-Center Study of 228 Consecutive Cases and Short-Term Outcomes.

Aims. We have established a procedure for uncut Roux-en-Y gastrojejunostomy after laparoscopic distal gastrectomy. This study aimed to evaluate the safety and technical feasibility of the procedure for patients with distal gastric cancer according to the short-term outcomes. Methods. Two hundred and twenty-eight consecutive patients who underwent a laparoscopic distal gastrectomy with uncut Roux-en-Y gastrojejunostomy from September 2014 to August 2018 were reviewed retrospectively. All the laparoscopic operations were performed successfully without conversion to open surgery. Results. The mean operative duration was 178.28 ± 32.82 minutes, the mean anastomotic process duration was 28.22 ± 7.50 minutes, the average blood loss was 48.97 ± 29.16 mL, and the overall number of lymph nodes harvested was 37.16 ± 11.47. The mean time of out-of-bed ambulation, anal exsufflation, liquid-diet intake, and duration of hospital stay were 41.99 ± 18.37 hours, 69.57 ± 23.17 hours, 5.06 ± 1.09 days, and 8.77 ± 2.42 days, respectively. Fifteen patients suffered postoperative complications, and the overall incidence rate was 6.58% (15/228). Seventeen patients experienced afferent recanalization, the mean time of which was 11 months after the operation. Conclusion. The laparoscopic uncut Roux-en-Y reconstruction is safe and technically feasible, and it has inspiring short-term outcomes for patients undergoing distal gastrectomy.

Increased ILC3s associated with higher levels of IL-1ß aggravates inflammatory arthritis in mice lacking phagocytic NADPH oxidase.

The role of redox regulation in immune-mediated arthritis has been previously described. However, the relationship between innate immune cells, including innate lymphoid cells (ILCs) and phagocyte-derived ROS, in this process remains unclear. Here, we characterize ILCs and measure the IL-1 family cytokines along with other cytokines relevant to ILC functions and development in serum-induced arthritic joints in wild type and phagocytic NADPH oxidase (NOX2)-deficient Ncf1-/- mice. We found more severe serum-induced joint inflammation and increased NCR+ ILC3s in inflamed joints of Ncf1-/- mice. Furthermore, in vitro stimulation with IL-1ß on Tbet+ ILC1s from joints facilitated their differentiation into ROR-γt+ ILC3s. Moreover, treatment with IL-1 antagonists effectively lowered the proportions of NCR+ ILC3s and IL-17A producing ILC3s in Ncf1-/- arthritic mice and ameliorated the joint inflammation. These results suggest that NOX2 is an essential regulator of ILC transdifferentiation and may mediate this process in a redox-dependent manner through IL-1ß production in the inflammatory joint. Our findings shed important light on the role of ILCs in the initiation and progression in tissue inflammation and delineate a novel innate immune cell-mediated pathogenic mechanism through which redox regulation may determine the direction of immune responses in joints.

Comparison of treatment outcomes between laparoscopic and endoscopic surgeries for relatively small gastric gastrointestinal stromal tumors.

BACKGROUND: Complete surgical resection remains the predominant treatment modality for primary gastrointestinal stromal tumors (GISTs). No therapeutic consensus exists for 2-5 cm gastric GISTs. We compared the efficacy, safety, and prognosis of laparoscopic and endoscopic surgeries in the treatment of relatively small (2-5 cm) intraluminal gastric GISTs. METHODS: We collected 101 patients with relatively small intraluminal gastric GISTs who had integrated clinicopathological data and underwent laparoscopic or endoscopic resection (laparoscopic group n = 66; endoscopic group n = 35). Clinicopathological characteristics, perioperative data, and long-term oncological outcomes were retrospectively analyzed. Comparative analysis of clinicopathological data in the two groups was performed by using a chi-square test, Fisher's exact test, and Student's t-test. Recurrence-free survival (RFS) was analyzed by the log-rank test. RESULTS: All clinicopathological characteristics had no significant difference between the two groups. Patients in the endoscopic group had shorter operation time (P < 0.001), postoperative hospital stay (P < 0.001), time to a liquid diet (P < 0.01), and time to a semi-liquid diet (P < 0.01), and lower hospital charges (P < 0.001), compared to those in the laparoscopic group. Four patients (6.1%) in the laparoscopic group and one patient (2.9%) in the endoscopic group had perioperative complications, but with no significant difference. Recurrence occurred in 6 patients (9.1%) and 2 patients (5.7%) in the laparoscopic and endoscopic groups, respectively. There was no significant difference in RFS between the two groups. CONCLUSION: Endoscopic resection is a feasible and safe treatment modality for patients with relatively small (2-5 cm) intraluminal gastric GISTs. Due to faster recovery and lower cost, endoscopic resection is more suitable for elderly and weak patients, or patients with a poor financial situation.

miR-324-3p promotes gastric cancer development by activating Smad4-mediated Wnt/beta-catenin signaling pathway.

BACKGROUND: Emerging evidence suggested that miRNAs can function as oncogenes or tumor suppressors by regulating downstream target genes. miR-324-3p has been reported to function in several carcinomas, but its role in gastric cancer (GC) is still unknown. This study aims to explore the effects of miR-324-3p on the development of GC. METHODS: Expression of miR-324-3p was examined in GC cells and tissues by qRT-PCR. Effects of miR-324-3p on GC cells were evaluated by cell vitality assay, colony formation assay, cell migration assay, and flow cytometric assay. The dual luciferase assay was used to verify whether miR-324-3p could interact with the potential target genes. Western blot was used to assess the expression level of Smad4 and beta-catenin. Intracellular ATP level was also examined. The tumor xenografts were established using nude mice. A gastric organoid model was made from fresh stomach tissue. RESULTS: miR-324-3p was expressed at higher levels in the tumor tissues compared with adjacent normal tissues. Overexpression of miR-324-3p promoted cell growth, migration, and decreased apoptosis. miR-324-3p repressed the expression of Smad4, and loss of Smad4 activated the Wnt/beta-catenin signaling pathway. Overexpression of Smad4 rescued the effects of miR-324-3p on GC cells. The intracellular ATP level was upregulated with overexpression of miR-324-3p. miR-324-3p facilitated tumor cell colonization and growth in vivo and contributed to the growth of gastric organoids. CONCLUSIONS: The results suggested that miR-324-3p promoted GC through activating the Smad4-mediated Wnt/beta-catenin signaling pathway. The miR-324-3p/Smad4/Wnt signaling axis may be a potential therapeutic target to prevent GC progression.

Natriuretic peptide receptor A inhibition suppresses gastric cancer development through reactive oxygen species-mediated G2/M cell cycle arrest and cell death.

Natriuretic peptide receptor A (NPRA), the major receptor for atrial natriuretic peptide (ANP), has been implicated in tumorigenesis; however, the role of ANP-NPRA signaling in the development of gastric cancer remains unclear. Immunohistochemical analyses indicated that NPRA expression was positively associated with gastric tumor size and cancer stage. NPRA inhibition by shRNA induced G2/M cell cycle arrest, cell death, and autophagy in gastric cancer cells, due to accumulation of reactive oxygen species (ROS). Either genetic or pharmacologic inhibition of autophagy led to caspase-dependent cell death. Therefore, autophagy induced by NPRA silencing may represent a cytoprotective mechanism. ROS accumulation activated c-Jun N-terminal kinase (JNK) and AMP-activated protein kinase (AMPK). ROS-mediated activation of JNK inhibited cell proliferation by disturbing cell cycle and decreased cell viability. In addition, AMPK activation promoted autophagy in NPRA-downregulated cancer cells. Overall, our results indicate that the inhibition of NPRA suppresses gastric cancer development and targeting NPRA may represent a promising strategy for the treatment of gastric cancer.

Redox Regulation of Pro-IL-1ß Processing May Contribute to the Increased Severity of Serum-Induced Arthritis in NOX2-Deficient Mice.

AIMS: To elucidate the role of reactive oxygen species (ROS) in arthritis and to identify targets of arthritis treatment in conditions with different levels of oxidant stress. RESULTS: Through establishing an arthritis model by injecting arthritogenic serum into wild-type and NADPH oxidase 2 (NOX2)-deficient mice, we found that arthritis had a neutrophilic infiltrate and was more severe in Ncf1(-/-) mice, a mouse strain lacking the expression of the NCF1/p47(phox) component of NOX2. The levels of interleukin-1ß (IL-1ß) and IL-6 in inflamed joints were higher in Ncf1(-/-) than in controls. Antagonists of tumor necrosis factor-α (TNFα) and IL-1ß were equally effective in suppressing arthritis in wild-type mice, while IL-1ß blockade was more effective than TNFα blockade in Ncf1(-/-) mice. A treatment of caspase inhibitor and the combination treatment of a caspase inhibitor and a cathepsin inhibitor, but not a cathepsin inhibitor alone, suppressed arthritic severity in the wild-type mice, while a treatment of cathepsin inhibitor and the combination treatment of a caspase inhibitor and a cathepsin inhibitor, but not a caspase inhibitor alone, were effective in treating Ncf1(-/-) mice. Consistently, cathepsin B was found to proteolytically process pro-IL-1ß to its active form and this activity was suppressed by ROS. INNOVATION: This novel mechanism of a redox-mediated immune regulation of arthritis through leukocyte-produced ROS is important for devising an optimal treatment for patients with different levels of tissue ROS. CONCLUSION: Our results suggest that ROS act as a negative feedback to constrain IL-1ß-mediated inflammation, accounting for the more severe arthritis in the absence of NOX2.

Secretory function in subplate neurons during cortical development.

Subplate cells are among the first generated neurons in the mammalian cerebral cortex and have been implicated in the establishment of cortical wiring. In rodents some subplate neurons persist into adulthood. Here we would like to highlight several converging findings which suggest a novel secretory function of subplate neurons during cortical development. Throughout the postnatal period in rodents, subplate neurons have highly developed rough endoplasmic reticulum (ER) and are under an ER stress condition. By comparing gene expression between subplate and layer 6, we found that several genes encoding secreted proteins are highly expressed in subplate neurons. One of these secreted proteins, neuroserpin, encoded by the serpini1 gene, is localized to the ER in subplate cells. We propose that subplate might influence cortical circuit formation through a transient secretory function.

Acute ethanol induces apoptosis by stimulating TRPC6 via elevation of superoxide in oxygenated podocytes.

Our recent studies indicate that hydrogen peroxide (H2O2) only at high concentrations can cause oxidative stress in renal epithelial cells and induce apoptosis of podocytes. Consistently, the present study shows that H2O2, even at 1 mM, failed to induce intracellular oxidative stress and apoptosis of the podocytes due to efficient activity of catalase, an enzyme which degrades H2O2 to produce water and oxygen (O2). However, H2O2 acted as a source of O2 to allow acute ethanol to induce superoxide production and cause apoptosis of the podocytes. In contrast, acute ethanol alone did not elevate intracellular superoxide, even though it stimulates expression and translocation of p47phox to the plasma membrane. Inhibition of catalase abolished not only O2 production from H2O2 degradation, but also NOX2-dependent superoxide production in the podocytes challenged by both H2O2 and acute ethanol. In parallel, acute ethanol in the presence of H2O2, but neither ethanol nor H2O2 alone, stimulated transient receptor potential canonical 6 (TRPC6) channels and caused TRPC6-dependent elevation of intracellular Ca2+. These data suggest that exogenous H2O2 does not induce oxidative stress due to rapid degradation to produce O2 in the podocytes, but the oxygenated podocytes become sensitive to acute ethanol challenge and undergo apoptosis via a TRPC6-dependent elevation of intracellular Ca2+. Since cultured podocytes are considered in hypoxic conditions, H2O2 may be used as a source of O2 to establish an ischemia-reperfusion model in some type of cultured cells in which H2O2 does not directly induce intracellular oxidative stress.

Lutein protects against ischemia/reperfusion injury in rat kidneys.

Ischemia­reperfusion (I/R) injury has a major impact on renal dysfunction during transplantation. The present study investigated the role of lutein against I/R injury­induced oxidative stress in rat kidneys. Biochemical analysis and oxidative stress parameters demonstrated that lutein protected the rat kidney significantly from I/R injury. Pretreatment with lutein significantly increased the total antioxidant capacity with a concomitant decline in the total oxidant status. Rats with I/R injury showed a significant increase in oxidative stress. The results revealed significant increases in the levels of lipid peroxidation and protein carbonyl content with concomitant decreases in enzymic and non­enzymic antioxidants. The activity of these enzymes was reversed and demonstrated a significant increase following lutein pre­treatment compared with the rats subjected to I/R injury alone. Furthermore, lutein protected the renal tissue from I/R injury by maintaining normal kidney architecture and led to a reduction in the levels of the renal markers urea and creatinine in the serum. These results demonstrated clear evidence that lutein offered a significant protective effect against I/R injury by enhancing antioxidant defense mechanisms.

Subplate in a rat model of preterm hypoxia-ischemia.

OBJECTIVE: Hypoxia-ischemia (HI) in preterm infants primarily leads to injuries in the cerebral white matter. However, there is growing evidence that perinatal injury in preterms can also involve other zones including the cortical gray matter. In a neonatal rat model of HI, selective vulnerability of subplate has been suggested using BrdU birth-dating methods. In this study, we aimed to investigate the neuropathological changes of the subplate and deep layers of the cortex following cerebral HI in neonatal rats with specific cell markers. METHODS: P2 rats underwent permanent occlusion of the right common carotid artery followed by a period of hypoxia. P8 rats were analyzed using immunohistochemistry; subplate and deep layers cells were quantified and compared with sham-operated case. RESULTS: A large variability in the extent of the cerebral injury was apparent. For the three analyzed subplate populations (Nurr1+, Cplx3+, and Ctgf+ cells), no significant cell reduction was observed in mild and moderate cases. Only in severe cases, subplate cells were strongly affected, but these injuries were always accompanied by the cell reductions in layers VI and V. INTERPRETATION: We could therefore not confirm a specific vulnerability of subplate cells compared to other deep layers or the white matter in our model.

Inverse associations of outdoor activity and vitamin D intake with the risk of Parkinson's disease.

Early studies had suggested that vitamin D intake was inversely associated with neurodegenerative diseases, such as Alzheimer's disease and multiple sclerosis. However, the associations of vitamin D intake and outdoor activities with Parkinson's disease (PD) are still unclear, so this study is to evaluate these relationships from a case-control study in elderly Chinese. The study population involved 209 cases with new onsets of PD and 210 controls without neurodegenerative diseases. The data on dietary vitamin D and outdoor activities were collected using a food-frequency questionnaire and self-report questionnaire. Multivariable logistic regressions were used to examine the associations between dietary outdoor activities, vitamin D intake and PD. Adjustment was made for sex, age, smoking, alcohol use, education, and body mass index (BMI). Adjusted odds ratios (ORs) for PD in quartiles for outdoor physical activity were 1 (reference), 0.739 (0.413, 1.321), 0.501 (0.282, 0.891), and 0.437 (0.241, 0.795), respectively (P=0.002 for trend). Adjusted ORs for PD in quartiles for total vitamin D intake were 1 (reference), 0.647 (0.357, 1.170), 0.571 (0.318, 1.022), and 0.538 (0.301, 0.960), respectively (P=0.011 for trend). Our study suggested that outdoor activity and total vitamin D intake were inversely associated with PD, and outdoor activity seems to be more significantly associated with decreased risk for PD.

Protective effect of cytosolic phospholipase A2 inhibition against inflammation and degeneration by promoting regulatory T cells in rats with experimental autoimmune encephalomyelitis.

Cytosolic phospholipase A2 (cPLA2) is the rate-limiting enzyme that initiates the production of various inflammatory mediators. Previous studies have shown that inhibiting cPLA2 exerts a neuroprotective effect on experimental autoimmune encephalomyelitis (EAE) by ameliorating the severity of the disease and influencing Th1 and Th17 responses. However, it remains unclear whether treatment with a cPLA2 inhibitor will influence the regulatory T cells (Tregs) that play a critical role in maintaining immune homeostasis and preventing autoimmune diseases. In this study, the cPLA2 inhibitor AX059 reduced the onset and progression of EAE in Lewis rats. In addition, this effect was accompanied by activation of Tregs and alterations in the expression of their various cytokines. The study therefore demonstrated that Tregs are involved in the immunomodulatory effect mediated by cPLA2 inhibition. These findings may have clinical application in the treatment of multiple sclerosis.

MicroRNA expression aberration in Chinese patients with relapsing remitting multiple sclerosis.

Multiple sclerosis (MS) is a chronic inflammatory demyelinating disease that is caused by an abnormal immune system that attacks the central nervous system. MicroRNAs (miRNAs) are known to play essential roles in the immune system. Most of the previous studies about miRNA dysregulation in MS have focused on European populations. In the present study, the miRNA expression profiles associated with Chinese MS patients are investigated. Here, human miRNA expression profiling experiments were performed on the peripheral blood mononuclear leukocytes from ten MS patients and ten healthy controls. Nine overexpressed and six underexpressed miRNAs were found. The 15 miRNAs were validated independently in a second cohort of 40 MS patients and 40 controls by real-time PCR; six miRNAs were significantly abnormally expressed, and principal component analysis of six miRNAs indicated that the MS patients could be clearly differentiated from the healthy controls based on the miRNA expression patterns. This study provided the indications of abnormal miRNA expression patterns in the peripheral blood mononuclear leukocytes of Chinese MS patients. The potential roles of these differentially expressed miRNAs as MS biomarker and in pathogenesis need to be investigated in future studies.

Adult pallium transcriptomes surprise in not reflecting predicted homologies across diverse chicken and mouse pallial sectors.

The thorniest problem in comparative neurobiology is the identification of the particular brain region of birds and reptiles that corresponds to the mammalian neocortex [Butler AB, Reiner A, Karten HJ (2011) Ann N Y Acad Sci 1225:14-27; Wang Y, Brzozowska-Prechtl A, Karten HJ (2010) Proc Natl Acad Sci USA 107(28):12676-12681]. We explored which genes are actively transcribed in the regions of controversial ancestry in a representative bird (chicken) and mammal (mouse) at adult stages. We conducted four analyses comparing the expression patterns of their 5,130 most highly expressed one-to-one orthologous genes that considered global patterns of expression specificity, strong gene markers, and coexpression networks. Our study demonstrates transcriptomic divergence, plausible convergence, and, in two exceptional cases, conservation between specialized avian and mammalian telencephalic regions. This large-scale study potentially resolves the complex relationship between developmental homology and functional characteristics on the molecular level and settles long-standing evolutionary debates.

Association between EXOC3L2 rs597668 polymorphism and Alzheimer's disease.

BACKGROUND: EXOC3L2 gene rs597668 polymorphism was identified to be significantly associated with Alzheimer's disease (AD) in Caucasian population. However, recent studies reported consistent and inconsistent results in Caucasian and Asian populations. AIMS: In order to assess this association, we performed a meta-analysis of rs597668 polymorphism using RevMan (v.5.1) software. METHODS: We searched PubMed and Google scholar databases and selected 4 independent publications, which included 16 independent studies. We conducted sensitivity analysis and evaluated the publication bias. In the end, we calculated the odds ratio (OR) using fixed effect model (Mantel-Haenszel). RESULTS: We observed significant association between rs597668 polymorphism and AD using allele model (P = 0.006, OR = 1.09, 95% CI 1.03-1.16) and the dominant model (P = 0.008, OR = 1.11, 95% CI 1.03-1.21). DISCUSSION AND CONCLUSIONS: To our knowledge, this is the first study that assesses the association between rs597668 polymorphism and AD by meta-analysis. We believe that our findings will be very useful for future genetic studies in AD.

Expression profiling of mouse subplate reveals a dynamic gene network and disease association with autism and schizophrenia.

The subplate zone is a highly dynamic transient sector of the developing cerebral cortex that contains some of the earliest generated neurons and the first functional synapses of the cerebral cortex. Subplate cells have important functions in early establishment and maturation of thalamocortical connections, as well as in the development of inhibitory cortical circuits in sensory areas. So far no role has been identified for cells in the subplate in the mature brain and disease association of the subplate-specific genes has not been analyzed systematically. Here we present gene expression evidence for distinct roles of the mouse subplate across development as well as unique molecular markers to extend the repertoire of subplate labels. Performing systematic comparisons between different ages (embryonic days 15 and 18, postnatal day 8, and adult), we reveal the dynamic and constant features of the markers labeling subplate cells during embryonic and early postnatal development and in the adult. This can be visualized using the online database of subplate gene expression at https://molnar.dpag.ox.ac.uk/subplate/. We also identify embryonic similarities in gene expression between the ventricular zones, intermediate zone, and subplate, and distinct postnatal similarities between subplate, layer 5, and layers 2/3. The genes expressed in a subplate-specific manner at some point during development show a statistically significant enrichment for association with autism spectrum disorders and schizophrenia. Our report emphasizes the importance of the study of transient features of the developing brain to better understand neurodevelopmental disorders.

Up-regulated GLT-1 resists glutamate toxicity and attenuates glutamate-induced calcium loading in cultured neurocytes.

Glutamate transporter-1 (GLT-1) plays a dual role in glutamate transportation: both normally devotion to the clearance of glutamate and during some pathological conditions extruding glutamate to the extracellular space. Therefore, it is uncertain whether increased expression of GLT-1 will actually be helpful against glutamate excitotoxicity. In this study, GLT-1 up-regulation was induced by ceftriaxone, and L-glutamate was added to induce glutamate toxicity in primary cultured rat cortical cells. The results showed that up-regulated GLT-1 induced by 1 µM ceftriaxone for 2 days markedly increased cell viability, decreased apoptotic cell death and alleviated ultrastructural damage induced by 50 µM glutamate 15 min. as well as promoted L-[(3) H]-glutamate uptake in cultured cells. GLT-1 up-regulation had no effect on the intracellular free calcium concentration ([Ca(2+) ](i) ) in the resting situation, while relieved intracellular calcium overloading by reducing the elevation and promoting the recovery of [Ca(2+) ](i) following stimulation of 50 µM glutamate for 2 min. Applying 100 µM dihydrokainic acid (GLT-1 antagonist) 30 sec. before glutamate eliminated the above effect of GLT-1 up-regulation on [Ca(2+) ](i) . In conclusion, GLT-1 up-regulation induced by ceftriaxone plays a positive glutamate transporting role against glutamate toxicity in primary cultured rat cortical cells.

Th17 helper cell and T-cell immunoglobulin and mucin domain 3 involvement in Guillain-Barré syndrome.

OBJECTIVE AND DESIGN: We investigated the involvement of Th17 cells and T-cell immunoglobulin and mucin domain 3 (TIM-3) in Guillain-Barré syndrome (GBS) in comparison to healthy subjects. MATERIALS AND SUBJECTS: Peripheral blood samples were obtained from 29 healthy subjects and 29 GBS patients. TREATMENT: Peripheral blood mononuclear cells (PBMCs) and CD4(+) T cells were stimulated with anti-CD3 and anti-CD28 mAbs, in the absence or presence of anti-TIM-3 mAb. METHODS: mRNA levels of TIM-3 and the transcription factor retinoic acid-related orphan receptor γt (RORγt) were determined by RT-PCR and were expressed relative to ß-actin mRNA (housekeeping gene). Serum IFN-γ and IL-17 levels were determined by ELISA. RESULTS: Compared to controls, relative TIM-3 mRNA levels were lower in both stimulated and unstimulated PBMCs from GBS patients. Unstimulated GBS CD4(+) T cells and GBS CD4+ T cells stimulated with anti-CD3 and CD28 mAbs had higher relative RORγt mRNA expression compared to controls. GBS CD4(+) T cells secreted significantly more IFN-γ and IL-17 in the presence of anti-TIM-3 mAb. GBS patients had (1) higher numbers of Th17, but not Th1 or Th2 cells in peripheral blood and (2) higher serum concentrations of IFN-γ and IL-17 compared to controls. CONCLUSION: TIM-3 may inhibit Th17 cell activation, thereby modulating their cytokine secretion patterns. Th17 cell differentiation, IL-17 levels, and TIM-3 regulation may be involved in the pathogenesis of GBS.