Unusual severe gastritis and gastric ulcers caused by pembrolizumab.

Pembrolizumab (an immune checkpoint inhibitor)-related gastritis and gastric ulcers are rare immune-related adverse events, which are insufficiently treated with proton pump inhibitors (PPIs) therapy alone, and usually require systemic steroid therapy and even other biological agents (such as infliximab) in severe cases. Here, we report a case of 49-years-old woman suffering from gastritis and gastric ulcers after pembrolizumab treatment, which was refractory to 2 months of PPI therapy. The diagnosis was made by the clinical and histopathologic presentations. She had immediate resolution of abdominal symptoms after initiation of steroid treatment, but the gastritis and gastric ulcers improved slowly and lasted for months as shown in endoscopy. She was finally treated with extended steroid therapy without serious complications. We discuss the latest treatment options and our management strategies of the case.

[Laparoscopic diagnosis of postoperative recurrence of peritoneal metastasis in gastric cancer patients and the clinical efficacy of bidirectional intraperitoneal and systemic chemotherapy].

Objective: To explore the diagnostic value of laparoscopy in the postoperative recurrence of peritoneal metastasis in gastric cancer, and to investigate the efficacy of bidirectional intraperitoneal and systemic (BIPS) chemotherapy for the recurrence. Methods: The descriptive case series study was conducted. Case inclusion criteria: (1) gastric cancer patients without synchronous distant metastasis received D2 radical gastrectomy; (2) postoperative adjuvant chemotherapy was administered; (3) no other distant metastasis except recurrence of peritoneal metastasis; (4) age of 18-75 years; (5) Eastern Cooperative Oncology Group (ECOG) performance-status score≤2; (6) pretreatment evaluation suggested that surgery and chemotherapy could be tolerated. Eight consecutive gastric cancer patients with postoperative recurrence of peritoneal metastasis who met the above criteria at Department of Gastrointestinal Surgery of Ruijin Hospital from September 2015 to September 2016 were enrolled into the study. There were 6 males and 2 females with the median age of 52 (38-68) years. They received laparoscopy or laparotomy first, and then were evaluated with reference to the Sugarbaker peritoneal cancer index (PCI) and the peritoneal metastasis classification of gastric cancer developed by the Japanese Gastric Cancer Research Association. A peritoneal access port was implanted in the subcutaneous space of the lower abdomen and the patients received chemotherapy for 21 days as a course of treatment. All the patients received intraperitoneal 20 mg/m(2) of paclitaxel (PTX) via implanted subcutaneous peritoneal access ports and intravenous 50 mg/m(2) of PTX at day 1 and day 8, meanwhile 80 mg/m(2) of Tigio was orally administered per day for 14 consecutive days, followed by 7 days of interval. Follow-up ended on December 15, 2019. Results: Of these 8 patients with recurrence of peritoneal metastasis after gastric cancer surgery, 1 case underwent laparotomy and loop stoma of terminal ileum because of complete colonic obstruction, and the remaining 7 cases underwent laparoscopy successfully and the recurrence of peritoneal metastasis was clearly diagnosed. Two patients with ovarian metastasis underwent laparoscopic bilateral adnexectomy. The median follow-up time was 17.5 (1.5 to 39.0) months, the median number of BIPS chemotherapy course was 11 (1 to 30), and the median survival time (MST) after BIPS chemotherapy was 17.0 months. The major adverse reaction in BIPS treatment was mainly myelosuppression, of which grade 3/4 leukopenia and neutropenia developed in 1 and 2 cases respectively. No BIPS-related death occurred. The MST of gastric cancer after radical gastrectomy was 40.0 months. Conclusions: Laparoscopy is a safe and feasible method for diagnosing the recurrence of peritoneal metastasis of gastric cancer. BIPS chemotherapy is effective and safe for its treatment and deserves further study.

[Clinical and genetic characteristics of children with STXBP1 encephalopathy].

Objective: To investigate the clinical and genetic characteristics of developmental and epileptic encephalopathy (DEE) caused by syntaxin-binding protein 1 (STXBP1) gene mutation. Methods: The clinical data, gene variation and treatment outcome of 15 children with STXBP1 encephalopathy admitted to Children's Hospital Affiliated to Capital Institute of Pediatrics from January 2014 to June 2019 were analyzed retrospectively. Results: Among 15 patients, 11 were male and 4 were female, age ranged from 2 months to 69 months. The clinical manifestations of 14 children were epilepsy and developmental delay (DD) and the remaining one showed developmental delay without seizure. The onset age of epilepsy ranged from two days to 19 months and 11 of them experienced the first attack before 1 year of age. The common seizure types were epileptic spasms and tonic seizures. Seven patients were diagnosed with Ohtahara syndrome or West syndrome. Epileptic form discharges were observed in the interictal electroencephalograms (EEG) of 11 patients, including multifocal discharges, suppression-burst and hypsarrhythmia. The brain magnetic resonance imaging of 7 children were abnormal, including myelin dysplasia, less white matter, lack of corpus callosum or hypoplasia. The follow-up time ranged from 2 months to 57 months, after the last follow-up, 3 cases were seizure free, 6 children showed partial response and the other 5 patients had no response on multitherapy. Six of 8 patients showed good responses to levetiracetam (LEV) monotherapy or in combination with other antiepileptic drugs (AEDs). Vigabatrin (VGB) was applied to 5 patients with epileptic spasms and 4 of them showed response. All patients showed different degrees of developmental delay while four of them showed autistic features. STXBP1 gene mutations were identified in all cases and there were 15 types of gene variations, including 8 missense mutations, 1 nonsense mutation, 5 frame shift mutations and 1 complex mutation. Five novel mutations were unreported before, including c.1193A>G, c.172delG, c.1769C>T, c.1038_1039delCC, c.348_351dupTGAA. Conclusions: Development delay and epilepsy are the major and independent clinical phenotypes in children with STXBP1 encephalopathy. The variation of STXBP1 gene is mainly de novo. Levetiracetam and vigabatrin may be more effective in epilepsy control than other AEDs.

[Analysis and comparison of the clinical features and prognosis between extra - gastrointestinal stromal tumors and duodenal gastrointestinal stromal tumors].

Objective: To investigate the differences of clinicopathological features, diagnosis, treatment and prognosis between patients with extra-gastrointestinal stromal tumors (EGIST) and duodenal gastrointestinal stromal tumors (DGIST). Methods: A retrospective case - control study was performed. Case inclusion criteria: (1) tumor confirmed by histology and pathology; (2) primary tumor locating in the extra - gastrointestinal tract or duodenum; (3) without other synchronous tumors; (4) complete clinical and pathological data. Clinical data of 20 EGIST patients and 32 DGIST patients from March 2011 to September 2016 at Department of Gastrointestinal Surgery, Ruijin Hospital, Shanghai Jiaotong University School of Medicine were retrospectively collected and analyzed. The observational parameters included clinicopathological characteristics, treatment and prognosis conditions. Continuous data of abnormal distribution were expressed as median (range) and compared using the Mann-Whitney U-test. Survival curves were drawn by the Kaplan-Meier method and compared with the Log-rank test. Results: Of the 20 EGIST patients, 8 were males and 12 were females with age of 61.0 (30.0 to 86.0) years and of the 32 DGIST patients, 12 were males and 20 were females with age of 55.5 (27.0 to 70.0) years. Compared with DGIST patients, EGIST patients were older (U=188.000, P=0.012], had larger tumor size [10.0 (3.0 to 29.0) cm vs. 4.0 (1.5 to 10.0) cm, U=98.500, P<0.001] and higher ratio of high risk classification [85.0% (17/20) vs. 12.5% (4/32), χ(2)=26.870, P<0.001]. Among the 20 EGIST patients, 5 were diagnosed with distal metastasis and received imatinib (400 mg/d), and the other 15 patients underwent radical resection who were included in survival analysis. All the 32 DGIST patients underwent radical resection. The median follow-up of whole group was 43 (14 to 76) months. The 3-year recurrence/metastasis-free survival rate of 15 cases undergoing radical resection in the EGIST group was 85.6%, which was lower than that of the DGIST group (88.6%), and the difference was not statistically significant (P=0.745). There was no significant difference in the 3-year overall survival rate between the EGIST group (92.9%) and the DGIST group (100%) (P=0.271). Conclusions: As compared to DGIST, EGIST mostly occurs in those with older age, larger tumor size and higher risk grade. The prognosis of EGIST patients after radical resection is similar to that of DGIST patients.

[Predictive factors for lymph node metastasis in patients with poorly differentiated early gastric cancer].

Objective: This study aimed to identify clinicopathological factors predictive of lymph node metastasis in patients with the poorly differentiated early gastric cancer (EGC) to assess the feasibility of using endoscopic submucosal dissection (ESD). Methods: The records of patients with poorly differentiated early gastric cancer undergoing gastric radical resection between January 2012 and December 2016 were reviewed in Ruijin hospital. Those with distant metastasis, two or more malignant tumors, remnant gastric cancer, neo adjuvant therapy, previous history of gastric surgery or clear history of perigastric lymphadenectomy, and mixed tumors were excluded. Age, sex, presence of ulcerous lesion, tumor size, tumor location, depth of invasion, type of differentiation, lymphatic vessel invasion, vascular invasion, nerve invasion and HER2 expression were collected. Univariate and multivariate stepwise logistic regression analyses were used to identify the independent risk factors of perigastric lymph node metastasis.According to the Guidelines for the Treatment of Gastric Cancer (2018 edition) of the Chinese Society of Clinical Oncology (CSCO), the expanded indications of ESD for EGC are as follows: (1)no ulcerative lesions, the maximum diameter of lesions >2 cm of differentiated intramucosal cancer; (2)ulcerative lesions, the maximum diameter of lesions ≤3 cm of differentiated intramucosal cancer; (3)no ulcerative lesions, undifferentiated intramucosal carcinoma with diameter ≤2 cm. The relationship between clinicopathological factors and lymph node metastasis was analyzed. Results: A total of 517 patients, aged 21-83 (57.1±11.7), including 307 males and 210 females, were enrolled in the study. Among them, 114 (22.0%) patients had lymph node metastasis. Univariate analysis showed that ulcerative lesion (P=0.042), tumor diameter (P=0.048), depth of invasion (P<0.001), location of tumors (P<0.001), lymphatic vessel invasion (P=0.009), vascular invasion (P<0.001) and nerve invasion (P=0.028) were related to lymph node metastasis after radical resection of poorly differentiated early adenocarcinoma. Age, sex, type of differentiation and HER2 expression were not significantly correlated to lymph node metastasis (P>0.05). Multivariate analysis showed that tumor size (OR=1.61, 95% CI: 1.03-2.52, P=0.037), depth of invasion (OR=2.77, 95% CI:1.66-4.63, P<0.001), lymphatic duct invasion (OR=14.74, 95% CI:1.58-137.36, P=0.018) were independent risk factors for lymph node metastasis in poorly differentiated EGC, and ulcerative lesion was not a risk factor for lymph node metastasis (OR=0.82, 95% CI:0.56-1.18,P=0.285). A total of 119 patients with poorly differentiated EGC fully complied with the relative indications of ESD recommended by the Japanese Statute and the criteria for radical resection after ESD. Among them, 14 (11.8%) still had perigastric lymph node metastasis, while the gender, tumor diameter, location, differentiation and HER2 expression were not associated with lymph node metastasis (P>0.05). Conclusion: For patients with poorly differentiated EGC, the application of ESD should be carefully weighed with precise assessment of tumor diameter, depth of invasion, and lymphatic duct invasion.

Microbial community composition and diversity in rice straw digestion bioreactors with and without dairy manure.

Anaerobic digestion (AD) uses a range of substrates to generate biogas, including energy crops such as globally abundant rice straw (RS). Unfortunately, RS is high in lignocellulosic material and has high to C:N ratios (~80:1), which makes it (alone) a comparatively poor substrate for AD. Co-digestion with dairy manure (DM) has been promoted as a method for balancing C:N ratios to improve RS AD whilst also treating another farm waste and co-producing a potentially useful fertiliser. However, past co-digestion studies have not directly compared RS AD microbial communities with and without DM additions, which has made it hard to assess all impacts of DM addition to RS AD processes. Here, four RS:DM ratios were contrasted in identical semi-continuous-fed AD bioreactors, and 100% RS was found to produce the highest specific methane yields (112 mL CH4/g VS/day; VS, volatile solids), which is over double yields achieved in the reactor with the highest DM content (30:70 RS:DM by mass; 48 mL CH4/g VS/day). To underpin these data, microbial communities were sequenced and characterised across the four reactors. Dominant operational taxonomic units (OTUs) in the 100% RS unit were Bacteroidetes/Firmicutes, whereas the 30:70 RS:DM unit was dominated by Proteobacteria/Spirochaetes, suggesting major microbial community shifts occur with DM additions. However, community richness was lowest with 100% RS (despite higher specific yields), suggesting particular OTUs may be more important to yields than microbial diversity. Further, ambient VFA and VS levels were significantly higher when no DM was added, suggesting DM-amended reactors may cope better with higher organic loading rates (OLR). Results show that RS AD without DM addition is feasible, although co-digestion with DM will probably allow higher OLRs, resulting in great RS throughput in farm AD units.

Aberrant expression of Rab1A and its prognostic significance in human colorectal cancer.

OBJECTIVE: Colorectal cancer (CRC) is one of the most common types of cancer worldwide. Emerging evidence has verified that Rab1A plays an oncogenic role in several human malignancies including breast cancer, lung cancer, and hepatocellular carcinomas. However, the clinical significance and prognostic impact of Rab1A in CRC is still unclear. PATIENTS AND METHODS: We initiated our investigation by immunohistochemistry and Western blot analysis to confirm Rab1A expression in CRC tissues. Meanwhile, the correlation of Rab1A expression and clinicopathologic features, as well as outcome in CRC patients, were retrospectively analyzed. RESULTS: In the issue, Rab1A is overexpressed in CRC tissues compared with matched noncancerous tissues. Meanwhile, high Rab1A expression was significantly associated with the TNM stage, lymph node metastasis, and peritoneal metastasis. In addition, multivariate analyses identified Rab1A expression and TNM stage as independent predictors for CRC patients. Moreover, Kaplan-Meier survival analysis showed that patients with high Rab1A expression had a significantly worse survival time than those with low Rab1A expression, which especially affected the survival in CRC patients with advanced stage. Spearman analysis suggested that there was a positive relationship between Rab1A expression and preoperative serum carcinoembryonic antigen (CEA) for CRC patients. CONCLUSIONS: These results suggested that Rab1A is an important diagnostic marker for CRC, and Rab1A can be used as a valuable biomarker for prognosis as well as peritoneal metastasis in CRC patients. Rab1A may prove to be clinically useful for developing a new therapeutic target of CRC treatment.

Paeoniflorin attenuates postoperative pain by suppressing Matrix Metalloproteinase-9/2 in mice.

BACKGROUND: Recent studies have implicated that matrix metalloproteinase (MMP)-9 and MMP-2 play key roles in neuropathic pain due to their facilitation of inflammatory cytokine maturation and induction of neuroinflammation. However, the role of MMP-9/2 in postoperative pain is still unclear. We previously suggested that the natural compound paeoniflorin inhibited microglia activation induced by morphine treatment. In the present study, we demonstrated that paeoniflorin could alleviate postoperative pain via specific inhibition of matrix metalloproteinases (MMPs). METHODS: Mice received a plantar incision surgery and their mechanical allodynia was assessed with von Frey filaments. The activity of MMP-9/2 was determined by gelatin zymography. Cell signalling was assayed by western blot and immunohistochemistry. RESULTS: The expression of MMP-9/2 was significantly increased in mice spinal cords with plantar incision surgery. Paeoniflorin remarkably suppressed the activity of MMP-9/2 and relieved plantar incision-induced mechanical allodynia. Interestingly, the administration of paeoniflorin blocked the maturation of interleukin-1ß, which is a critical substrate of MMPs. Thereafter, paeoniflorin markedly suppressed microglia activation, inhibited the phosphorylation of p38 mitogen-activated protein kinase (MAPK) and the expression of neuronal c-Fos. CONCLUSION: These results indicated that MMP-9/2 activation in spinal microglia plays a key role in incision-induced mechanical allodynia in mice. Moreover, utilizing paeniflorin blockage of the microglia MMP-9/2 activity might represent a valuable alternative for treating postoperative pain. SIGNIFICANCE: Our results provided direct evidence for the first time that paeoniflorin can inhibit plantar incision-induced microglia TLR4/MMP-9/2/IL-1ß signalling pathway and suppress postoperative pain. Thus, regulation of microglia MMP-9/2 may provide a new strategy for ameliorating postoperative pain.

[Clinical features of autoimmune hepatitis patients with poor response to treatment].

Objective: To investigate the clinical features of autoimmune hepatitis (AIH) patients with poor response to treatment. Methods: A total of 61 AIH patients were enrolled, among whom 49 (80.33%) achieved complete response (good response group) and 12 (19.67%) had incomplete response (poor response group). The two groups were compared in terms of clinical manifestations, laboratory markers, abdominal ultrasound findings, pathological features by liver biopsy, and response to treatment. Continuous data were expressed as mean ± standard deviation (x±s), and the t-test was used for comparison between groups; categorical data were expressed as rates or percentages, and the chi-square test was used for comparison between groups; a binary logistic regression analysis was used to determine influencing factors. Results: Most patients were female in both groups, and there were no significant differences in sex ratio, mean age of onset, and general status including extrahepatic autoimmune disease between the two groups. Compared with the good response group, the poor response group had significantly higher levels of alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase (ALP), total bilirubin, immunoglobulin G, and immunoglobulin M (P < 0.05). Compared with the good response group, the poor response group had a significantly higher positive rate of autoimmune antibodies except anti-nuclear antibody (ANA), anti-smooth muscle antibody (SMA), antimitochondrial antibody (AMA), and AMA/M2 (75% vs 16.3%, P < 0.001), and there was a significant difference in the positive rate of gp210 antibody between the two groups (25% vs 0%, P < 0.01). There were significant differences between the poor response group and the good response group in the proportion of patients with liver cirrhosis (50.0 % vs 16.3%, P < 0.05) and splenomegaly (58.3% vs 22.4%, P < 0.05). The binary logistic regression analysis showed that a high serum level of ALP (odds ratio [OR] = 1.017, 95% confidence interval [CI] 1.001-1.033, P = 0.034), positive autoimmune antibodies except ANA, SMA, and AMA/M2 (OR = 70.842, 95% CI 2.132-2 354.371, P = 0.017), and liver cirrhosis (OR = 28.777, 95% CI 1.015-815.854, P = 0.049) were independent risk factors for initial treatment outcome. Conclusion: A high serum level of ALP, positive autoimmune antibodies except ANA, SMA, and AMA/M2, and liver cirrhosis are closely associated with poor response in AIH patients.

TGF-ß upregulates the translation of USP15 via the PI3K/AKT pathway to promote p53 stability.

Crosstalk between transforming growth factor beta (TGF-ß) signaling and p53 has a critical role in cancer progression. TGF-ß signals via Smad and non-Smad pathways. Under normal conditions, wild-type p53 forms a complex with Smad2/3 and co-activates transcription of a variety of tumor suppressor genes, resulting in tumor suppressive effects. Thus, p53 stability is essential in progression of tumor suppressive responses mediated by TGF-ß signaling. However, it remains unknown whether p53 stability is regulated by TGF-ß. In the current study, we identify that USP15 binds to and stabilizes p53 through deubiquitination in U2OS and HEK293 cells. TGF-ß promotes the translation of USP15 through activation of mammalian target of rapamycin by the phosphoinositide 3-kinase/AKT pathway. Upregulation of USP15 translation links the crosstalk between TGF-ß signaling and p53 stability, allowing this cytokine to have a critical role in cancer progression.

Seasonal concentrations, contamination levels, and health risk assessment of arsenic and heavy metals in the suspended particulate matter from an urban household environment in a metropolitan city, Beijing, China.

The levels and health risks of arsenic and heavy metals (As, Ba, Cd, Co, Cr, Cu, Mn, Ni, Pb, and Zn) in the suspended particulate matter (SPM) collected from an urban household environment in Beijing of China for 12 months were investigated. The mean concentrations of the studied toxic elements were higher and lower than crustal abundance and PM2.5 in the urban outdoors of Beijing. The concentrations of the studied elements displayed significant seasonality. The highest concentrations of the total elements occurred in winter, followed by autumn, while the lowest concentrations were recorded in summer. Based on the calculated values of enrichment factor (EF) and geoaccumulation index (Igeo), the levels for As and Cu were heavily contaminated, while those for Cd, Pb, and Zn were extremely contaminated. As and Pb might pose risks to children and adults via ingestion exposure. The accumulative risks of multi-elements resulted from dermal contact and inhalation exposures were not negligible. More attention should be paid to reducing the non-carcinogenic and carcinogenic health risks posed by the toxic elements bound to urban household SPM particles via ingestion, inhalation, and dermal contact exposure.

Pyrosequencing reveals bacterial communities in unchlorinated drinking water distribution system: an integral study of bulk water, suspended solids, loose deposits, and pipe wall biofilm.

The current understanding of drinking water distribution system (DWDS) microbiology is limited to pipe wall biofilm and bulk water; the contributions of particle-associated bacteria (from suspended solids and loose deposits) have long been neglected. Analyzing the composition and correlation of bacterial communities from different phases helped us to locate where most of the bacteria are and understand the interactions among these phases. In the present study, the bacteria from four critical phases of an unchlorinated DWDS, including bulk water, pipe wall biofilm, suspended solids, and loose deposits, were quantified and identified by adenosine triphosphate analysis and pyrosequencing, respectively. The results showed that the bulk water bacteria (including the contribution of suspended solids) contributed less than 2% of the total bacteria. The bacteria associated with loose deposits and pipe wall biofilm that accumulated in the DWDS accounted for over 98% of the total bacteria, and the contributions of bacteria in loose deposits and pipe wall biofilm were comparable. Depending on the amount of loose deposits, its contribution can be 7-fold higher than the pipe wall biofilm. Pyrosequencing revealed relatively stable bacterial communities in bulk water, pipe wall biofilm, and suspended solids throughout the distribution system; however, the communities present in loose deposits were dependent on the amount of loose deposits locally. Bacteria within the phases of suspended solids, loose deposits, and pipe wall biofilm were similar in phylogenetic composition. The bulk water bacteria (dominated by Polaromonas spp.) were clearly different from the bacteria from the other three phases (dominated by Sphingomonas spp.). This study highlighted that the integral DWDS ecology should include contributions from all of the four phases, especially the bacteria harbored by loose deposits. The accumulation of loose deposits and the aging process create variable microenvironments inside loose deposits structures for bacteria to grow. Moreover, loose deposits protect the associated bacteria from disinfectants, and due to their mobility, the associated bacteria reach taps easily.

Prophylactic thyroidectomy for MEN 2-related medullary thyroid carcinoma based on predictive testing for RET proto-oncogene mutation and basal serum calcitonin in China.

INTRODUCTION: Early and normative surgery is the only curative method for multiple endocrine neoplasia type 2 (MEN 2)-related medullary thyroid carcinoma (MTC). AIMS: To study the timing of prophylactic total thyroidectomy (TT) for MEN 2-related MTC with different RET mutations in a Chinese population, and to compare the sensitivity and accuracy of fully-automated chemiluminescence immunoassay (FACLIA) and radioimmunoassay (RIA) for serum calcitonin (Ct). METHODS: We collected 24 asymptomatic individuals from 8 unrelated Chinese families with MEN 2, and analyzed RET mutation and Ct levels. Then we performed TT on 17 of the 24 individuals, including TT (2/17), TT with bilateral level VI lymph-node dissection (B-LND(VI); 12/17) and TT with B-LND(VI) + modified unilateral/bilateral/local neck dissection (3/17). RESULTS: Histopathology revealed bilateral/unilateral MTC in 15/17 (88.2%; median diameter, 1.0 cm) and bilateral C-cell hyperplasia in 2/17 (11.8%; p.V292M/R67H/R982C and p.C618Y). Lymph-node metastasis/fibro-adipose tissue invasion (p.C634R) or solely fibro-adipose tissue invasion (p.C634Y) were found in 2/17 (11.8%). Elevated pre-surgical Ct (pre-Ct) was identified by FACLIA in 17/17 (median age, 24.0), while pre-Ct by RIA was found in only 6/15 (P < 0.001). The median follow-up was 22.0 months, during which 16/17 had no abnormality (one p.C634R individual had elevated Ct), and another 7 carriers still had consistently undetectable Ct by FACLIA. CONCLUSIONS: Our study highlights the importance and feasibility of individualized prophylactic TT for MEN 2-related MTC, based on predictive integrated screening of RET and pre-Ct levels. Besides, we recommend FACLIA to measure Ct for earlier diagnosis, treatment and follow-up monitoring of MTC.

Quantification and identification of particle-associated bacteria in unchlorinated drinking water from three treatment plants by cultivation-independent methods.

Water quality regulations commonly place quantitative limits on the number of organisms (e.g., heterotrophic plate count and coliforms) without considering the presence of multiple cells per particle, which is only counted as one regardless how many cells attached. Therefore, it is important to quantify particle-associated bacteria (PAB), especially cells per particle. In addition, PAB may house (opportunistic) pathogens and have higher resistance to disinfection than planktonic bacteria. It is essential to know bacterial distribution on particles. However, limited information is available on quantification and identification of PAB in drinking water. In the present study, PAB were sampled from the unchlorinated drinking water at three treatment plants in the Netherlands, each with different particle compositions. Adenosine triphosphate (ATP) and total cell counts (TCC) with flow cytometry were used to quantify the PAB, and high-throughput pyrosequencing was used to identify them. The number and activity of PAB ranged from 1.0 to 3.5 × 10(3) cells ml(-1) and 0.04-0.154 ng l(-1) ATP. There were between 25 and 50 cells found to be attached on a single particle. ATP per cell in PAB was higher than in planktonic bacteria. Among the identified sequences, Proteobacteria were found to be the most dominant phylum at all locations, followed by OP3 candidate division and Nitrospirae. Sequences related to anoxic bacteria from the OP3 candidate division and other anaerobic bacteria were detected. Genera of bacteria were found appear to be consistent with the major element composition of the associated particles. The presence of multiple cells per particle challenges the use of quantitative methods such as HPC and Coliforms that are used in the current drinking water quality regulations. The detection of anoxic and anaerobic bacteria suggests the ecological importance of PAB in drinking water distribution systems.

Hyperbaric oxygen therapy attenuates neuropathic hyperalgesia in rats and idiopathic trigeminal neuralgia in patients.

BACKGROUND: Neuropathic pain after nerve injury is severe and intractable, and current drug and non-drug therapies offer very limited pain relief. Hyperbaric oxygen (HBO 2) has been clinically used for protection of the nervous system after acute injury. We investigated whether HBO 2 treatment could prevent and/or attenuate neuropathic pain in animals and in patients. METHODS: Mechanical allodynia and thermal hyperalgesia and neurochemical alterations of neuropathic pain were analysed in male, adult, Sprague-Dawley rats with sciatic nerve injury. Clinical trials were conducted in patients with idiopathic trigeminal neuralgia. RESULTS: Repetitive HBO 2 treatment [a combination of pressure at 3 atmosphere absolute (ATA) and pure oxygen] greatly inhibited behavioural signs of neuropathic pain manifested as thermal hyperalgesia and mechanical allodynia. Such an HBO 2 treatment also inhibited nerve injury-induced induction of c-Fos and activation of astrocytes and increased phosphorylation of NR2B receptor and the subsequent Ca 2+-dependent signals in rats. Neither high pressure (up to 3 ATA) nor pure oxygen alone resulted in analgesic effect. In clinical trials, one course of HBO 2 therapy (10 consecutive days) produced a rapid-onset, dose-dependent and long-lasting analgesic effects evidenced by the decreased doses of carbamazepine required for keeping patient pain at a minimum and decreased scores of visual analogue scales, which was used for patient's self-evaluation. CONCLUSIONS: These findings support that HBO 2 therapy is an effective approach for treating neuropathic pain in both animals and human beings and suggest that neural protection, anti-inflammation and inhibition of nerve injury-induced altered neural activity may contribute to the analgesic effect of HBO 2 therapy.

Case report: a p.C618S RET proto-oncogene germline mutation in a large Chinese pedigree with familial medullary thyroid carcinoma.

We report a Chinese pedigree with familial medullary thyroid carcinoma. Direct sequencing of the entire coding sequences of Rearranged during Transfection (RET) identified a recurrent c.T1852A (p.C618S) mutation in 13 of 23 members. The polymorphisms c.A135G (p.A45A), c.A1296G (p.A432A), c.T2307G (p.L769L) and IVS19 + 15T > C were also found in 13 carriers, and c.G2073A (p.G691S) was found in 1 carrier. Of the 13 carriers, seven (mean age: 42.6 years, range: 27-64) presented MTC as the isolated clinical phenotype, with elevated basal serum calcitonin (average: 1077.9 ng/L, range: 504-2,652) and a mean diameter of thyroid nodules of 2.97 cm (range: 1.6-4.3); they underwent a total thyroidectomy with modified bilateral/unilateral neck dissection and/or level VI lymph node dissection. The other 6 carriers did not accept surgery (4 rejected, 2 awaited). These were 2 older patients (63 and 32 years) with elevated calcitonin (1359 and 41.4 ng/L) and multi-centric hypoechoic nodules (1.5 and 0.6 cm) with calcifications in both/left thyroid lobes; and Doppler ultrasound showed normal bilateral thyroids in 4 younger carriers (median age: 8.3 years, range: 4-12) but with increased calcitonin (average: 9.7 ng/L, range: 7.87-12.2) in 3 of them. The phenotype here is consistent with the clinical symptoms reported worldwide. We recommend that screening of hotspot regions of RET should be preferentially carried out, while whole-exon sequencing should be performed when clinical signs fail to reveal hotspot mutations or different phenotype discrepancies. Moreover, we strongly suggest prophylactic thyroidectomy should be performed before age 5 in carriers with p.C618S to prevent the occurrence and metastasis of MTC.

Apoptosis-dependent and -independent mechanisms mediate the phagocytic recognition/clearance of the HL60-A1 transfectants with prolonged survival.

OBJECTIVE: The phagocytic recognition and clearance of the recruited inflammatory cells with prolonged survival play a pivotal role in relieving tissue inflammation and maintaining tissue homeostasis. Transgenic mice expressing Bcl-2 in mature neutrophils demonstrated that Bcl-2 attenuated neutrophil apoptosis, while the homeostasis of the neutrophil population was essentially unaffected. This result suggests that clearance of neutrophils with prolonged survival operates independently from apoptosis. Owing to the constitutive and inducible expression of Bcl-2 homologue, A1 in human neutrophils and the intolerance of preparation for the isolated human neutrophils with prolonged survival, the human promyelocytic HL60-A1 transfectants were established to study the mechanism of phagocytic recognition/clearance of the cells with prolonged survival. MATERIALS AND METHODS: The non-apoptotic cells with prolonged survival were enriched by serum withdrawal for five days and negatively isolated by annexin V-binding beads. Then, the cells were labeled with a fluorogenic marker. Monocyte-derived macrophages (MDM) were co-cultured to perform the phagocytosis assay, and flow cytometry was employed to determine the phagocytic index. RESULTS: In the serum-free condition, the phagocytic index of HL60-A1 transfectants was little different from that of the HL60-EGFP control, despite showing a significantly lower degree of apoptosis. While the phagocytic index of HL60-EGFP control was significantly correlated with the degree of apoptosis, the index of the HL60-A1 transfectants was less relevant to it. The phagocytic index for the annexin V-positive cells did not distinguish the two cell types. However, the phagocytic index for the annexin V-negative cells from the HL60-A1 transfectants was increased with age in days. Preincubation of MDM with the scavenger receptor inhibitor, Oxi-LDL, and the inhibitory antibodies against alphavbeta3, CD14 and CD36 surface molecules could attenuate the phagocytic recognition of the annexin V-positive HL60 cells but not the annexin V-negative A1 transfectants with prolonged survival. CONCLUSIONS: This study thus suggests that a mechanism unrelated to apoptosis exists, which mediates the phagocytic clearance of the non-apoptotic cells with prolonged survival and may be associated with A1 function in the myeloid cells.

Proliferation of glycogen accumulating organisms induced by Fe(III) dosing in a domestic wastewater treatment plant.

To meet the effluent requirements given for the sensitive receiving body, the Southpest Wastewater Treatment Plant of Budapest, Hungary uses a combined activated sludge-biofilter system with chemical precipitation for P removal. Causes of the proliferation of glycogen accumulating organisms (GAOs) observed in the unaerated/oxic activated sludge unit of this system were investigated both in full-scale and in lab-scale experiments combined with a detailed analysis of the microbial communities. Concentration profile measurements throughout the 8-stage activated sludge unit indicated anaerobic conditions in the first two unaerated reactors and low orthophosphate level (< 1 mg l(-1)) in all of the stages that could not be attributed to the influent quality, but to Fe (III)-dosing to the returned activated sludge. Microbiological analysis revealed the presence of GAOs from the GB group in the Gammaproteobacteria and occasionally tetrad-forming organisms from Actinobacteria, and the absence of Rhodocyclus-related polyphosphate accumulating organisms (PAOs) in the activated sludge samples. Comparative lab-scale studies carried out in two identically arranged UCT-systems with staged anoxic reactors also confirmed that Fe (III)-dosing may result in phosphorus deficiency of the microbial niche, leading to the suppression of growth and EBPR activity of PAOs and to the proliferation of GAOs.

Community structure of microbial biofilms associated with membrane-based water purification processes as revealed using a polyphasic approach.

The microbial communities of membrane biofilms occurring in two full-scale water purification processes employing microfiltration (MF) and reverse osmosis (RO) membranes were characterized using a polyphasic approach that employed bacterial cultivation, 16S rDNA clone library and fluorescence in situ hybridization techniques. All methods showed that the alpha-Proteobacteria was the largest microbial fraction in the samples, followed by the gamma-Proteobacteria. This suggested that members of these two groups could be responsible for the biofouling on the membranes studied. Furthermore, the microbial community structures between the MF and RO samples were considerably different in composition of the most predominant 16S rDNA clones and bacterial isolates from the alpha-Proteobacteria and only shared two common groups ( Bradyrhizobium, Bosea) out of more than 17 different bacterial groups observed. The MF and RO samples further contained Planctomycetes and Fibroacter/ Acidobacteria as the second predominant bacterial clones, respectively, and differed in minor bacterial clones and isolates. The community structure differences were mainly attributed to differences in feed water, process configurations and operating environments, such as the pressure and hydrodynamic conditions present in the water purification systems.