Comprehensive assessment of HF-rTMS treatment mechanism for post-stroke dysphagia in rats by integration of fecal metabolomics and 16S rRNA sequencing.

Background: The mechanism by which high-frequency repetitive transcranial magnetic stimulation (HF-rTMS) improves swallowing function by regulating intestinal flora remains unexplored. We aimed to evaluate this using fecal metabolomics and 16S rRNA sequencing. Methods: A Post-stroke dysphagia (PSD) rat model was established by middle cerebral artery occlusion. The magnetic stimulation group received HF-rTMS from the 7th day post-operation up to 14th day post-surgery. Swallowing function was assessed using a videofluoroscopic swallowing study (VFSS). Hematoxylin-eosin (H&E) staining was used to assess histopathological changes in the intestinal tissue. Intestinal flora levels were evaluated by sequencing the 16S rRNA V3-V4 region. Metabolite changes within the intestinal flora were evaluated by fecal metabolomics using liquid chromatography-tandem mass spectrometry. Results: VFSS showed that the bolus area and pharyngeal bolus speed were significantly decreased in PSD rats, while the bolus area increased and pharyngeal transit time decreased after HF-rTMS administration (p < 0.05). In the PSD groups, H&E staining revealed damaged surface epithelial cells and disrupted cryptal glands, whereas HF-rTMS reinforced the integrity of the intestinal epithelial cells. 16S rRNA sequencing indicated that PSD can disturb the intestinal flora and its associated metabolites, whereas HF-rTMS can significantly regulate the composition of the intestinal microflora. Firmicutes and Lactobacillus abundances were lower in the PSD group than in the baseline group at the phylum and genus levels, respectively; however, both increased after HF-rTMS administration. Levels of ceramides (Cer), free fatty acids (FA), phosphatidylethanolamine (PE), triacylglycerol (TAG), and sulfoquinovosyl diacylglycerol were increased in the PSD group. The Cer, FA, and DG levels decreased after HF-rTMS treatment, whereas the TAG levels increased. Peptococcaceae was negatively correlated with Cer, Streptococcus was negatively correlated with DG, and Acutalibacter was positively correlated with FA and Cer. However, these changes were effectively restored by HF-rTMS, resulting in recovery from dysphagia. Conclusion: These findings suggest a synergistic role for the gut microbiota and fecal metabolites in the development of PSD and the therapeutic mechanisms underlying HF-rTMS.

Serum proteome and metabolome uncover novel biomarkers for the assessment of disease activity and diagnosing of systemic lupus erythematosus.

Systemic lupus erythematosus (SLE) is an autoimmune disease affecting thousands of people. There are still no effective biomarkers for SLE diagnosis and disease activity assessment. We performed proteomics and metabolomics analyses of serum from 121 SLE patients and 106 healthy individuals, and identified 90 proteins and 76 metabolites significantly changed. Several apolipoproteins and the metabolite arachidonic acid were significantly associated with disease activity. Apolipoprotein A-IV (APOA4), LysoPC(16:0), punicic acid and stearidonic acid were correlated with renal function. Random forest model using the significantly changed molecules identified 3 proteins including ATRN, THBS1 and SERPINC1, and 5 metabolites including cholesterol, palmitoleoylethanolamide, octadecanamide, palmitamide and linoleoylethanolamide, as potential biomarkers for SLE diagnosis. Those biomarkers were further validated in an independent cohort with high accuracy (AUC = 0.862 and 0.898 for protein and metabolite biomarkers respectively). This unbiased screening has led to the discovery of novel molecules for SLE disease activity assessment and SLE classification.

Assessment of variation in B-cell receptor heavy chain repertoire in patients with end-stage renal disease by high-throughput sequencing.

Background/Aims: End-stage renal disease (ESRD), characterized by progressive loss of rental function during the disease course, has been reported to be correlated with immune dysregulation. To date, a majority of previous studies on immune response to ESRD have been focused on the T-cell response. This prospective study was to assess the B-cell receptor (BCR) heavy chain repertoire in ESRD patients.Materials and methods: A total of 10 ESRD patients and six healthy controls were prospectively enrolled in this study. BCR immunoglobulin heavy chain (IGH) repertoire in the peripheral blood from ESRD patients and healthy individuals were analyzed by means of next generation sequencing (NGS) in combination with multiplex PCR, Illumina sequencing, and the international ImMunoGeneTics database (IMGT).Results: Abnormal BCR complementary-determining region 3 (CDR3) sequences were identified in relation to ESRD. We also found that the degree of the B-cell clonal expansion in the ESRD group was significantly greater than that in the control group (p < .05), whereas the distributions of BCR CDR3, V, D, J, and V-J gene segments were comparable between the ESRD and control groups. T-test for analysis of the distribution ratio of the V, D, J, and V-J genes revealed five up-regulated genes and nine down-regulated genes associated with ESRD, and there were significant differences between the ESRD and control groups (p < .05).Conclusions: We have provided a successful approach to analyzing peripheral B-cell repertoire in ESRD patients, and the results suggest a direct correlation between the BCR repertoire and ESRD. The ESRD-specific BCR CDR3 sequences may hold promise for potentially therapeutic benefit.

Development of a novel and economical agar-based non-adherent three-dimensional culture method for enrichment of cancer stem-like cells.

BACKGROUND: Non-adherent or ultra-low attachment three-dimensional (3D) culture, also called sphere formation assay, has been widely used to assess the malignant phenotype and stemness potential of transformed or cancer cells. This method is also popularly used to isolate the cancer stem-like cells (CSCs) or tumor-initiating cells based on their unique anchorage-independent growth or anoikis-resistant capacity. Different non-adhesive coating agents, such as poly-2-hydroxyethyl methacrylate (poly-HEMA) and synthetic hydrogels, have been used in this non-adherent 3D culture. However, preparation of non-adherent culture-ware is labor-intensive and technically demanding, and also costs of commercial non-adherent culture-ware prepared with various coating agents are relatively expensive and the culture-ware cannot be used repeatedly. METHODS: In this study, we developed a non-adherent 3D culture method based on agar coating for growing tumor spheres derived from various cancer cell lines and primary prostate cancer tissues under a non-adherent and serum-free condition. The tumor spheres generated by this 3D culture method were analyzed on their expression profiles of CSC-associated markers by reverse transcription quantitative polymerase chain reaction, presence and relative proportion of CSCs by fluorescence-activated cell sorting (CD133+/CD44+ cell sorting) and also a CSC-visualizing reporter system responsive to OCT4 and SOX2 (SORE6), and in vivo tumorigenicity. The repeated use of agar-coated plates for serial passages of tumor spheres was also evaluated. RESULTS: Our results validated that the multicellular tumor spheres generated by this culture method were enriched of CSCs, as evidenced by their enhanced expression profiles of CSC markers, presence of CD133+/CD44+ or SORE6+ cells, enhanced self-renewal capacity, and in vivo tumorigenicity, indicating its usefulness in isolation and enrichment of CSCs. The agar-coated plates could be used multiple times in serial passages of tumor spheres. CONCLUSIONS: The described agar-based 3D culture method offers several advantages as compared with other methods in isolation of CSCs, including its simplicity and low-cost and repeated use of agar-coated plates for continuous passages of CSC-enriched spheres.

Application of modified primary closure of the pelvic floor in laparoscopic extralevator abdominal perineal excision for low rectal cancer.

AIM: To introduce a novel, modified primary closure technique of laparoscopic extralevator abdominal perineal excision (LELAPE) for low rectal cancer. METHODS: We retrospectively analyzed data from 76 patients with rectal cancer who underwent LELAPE from March 2013 to May 2016. Patients were classified into the modified primary closure group (32 patients) and the biological mesh closure group (44 patients). The total operating time, reconstruction time, postoperative stay duration, total cost, postoperative complications and tumor recurrence were compared. RESULTS: All surgery was successfully performed. The pelvic reconstruction time was 14.6 ± 3.7 min for the modified primary closure group, which was significantly longer than that of the biological mesh closure group (7.2 ± 1.9 min, P < 0.001). The total operating time was not different between the two groups (236 ± 20 min vs 248 ± 43 min, P = 0.143). The postoperative hospital stay duration was 8.1 ± 1.9 d, and the total cost was 9297 ± 1260 USD for the modified primary closure group. Notably, both of these categories were significantly lower in this group than those of the biological mesh closure group (P = 0.001 and P = 0.003, respectively). There were no differences observed between groups when comparing other perioperative data, long-term complications or oncological outcomes. CONCLUSION: The modified primary closure method for reconstruction of the pelvic floor in LELAPE for low rectal cancer is technically feasible, safe and cost-effective.

Decolorization of azo dye C.I. Reactive Black 5 by ozonation in aqueous solution: influencing factors, degradation products, reaction pathway and toxicity assessment.

In this study, ozonation treatment of C.I. Reactive Black 5 (RB5) was investigated at various operating parameters. The results showed that the aqueous solution initially containing 200 mg/L RB5 was quickly decolorized at pH 8.0 with an ozone dose of 3.2 g/h. Reaction intermediates with m/z 281, 546, 201, 350, 286 and 222 were elucidated using liquid chromatography-mass spectrometry, while sulfate ion, nitrate ion and three carboxylic acids (i.e., oxalic acid, formic acid, and acetic acid) were identified by ion exchange chromatography. Thus, the cleavage of the azo bond and the introduction of OH groups in the corresponding positions were proposed as the predominant reaction pathway. The detachment of sulfonic groups was also commonly observed during the ozonation treatment. The proposed degradation mechanism was confirmed by frontier electron density calculations, suggesting the feasibility of predicting the major events in the whole ozonation process with the computational method. Compared with RB5 degradation, the reduction of total organic carbon (TOC) proceeded much more slowly, and approximately 54% TOC was removed after 4 h of ozonation. Acute toxicity tests with Photobacterium phosphoreum showed that the toxicity of reaction solution was firstly increased and then decreased to a negligible level after 160 min.

[Assessment of rice achievable productivity and its application in rice production management division: a case study in Fuyang County of Zhejiang Province, East China].

Rice productivity is the key factor affecting rice production and its sustainable development. Based on the gradation of cultivated land quality at county-level, this paper evaluated the rice productivity in Fuyang County of Zhejiang Province, and, through selected sampling field investigation and according to the land productivity index of paddy field quality, a model for assessing rice achievable productivity was established, aimed to analyze the regional rice productivity and its achievable productivity. In the study area, there was a positive correlation between the land productivity index and rice yield. For single cropping rice, its achievable productivity was 1.70 x 10(5) t, being 1.6 times of its realistic productivity (1.04 x 10(5) t). In 2009, the realistic rice productivity per unit area was 7676 kg x hm(-2), and the achievable productivity was 8831 kg x hm(-2), with a production potentiality of +15%, a big potential of rice production capacity in the county. Through the analyses of rice productivity, relative superiority of rice production scale, and its growth potential index in the villages and towns, the rice production of Fuyang County was divided into three regions, i. e., key enhancement region, optimization construction region, and development protection region.

[Assessment research on the inter-proximal mesial papilla height of single implant-supported maxillary central incisor].

OBJECTIVE: This prospective study was to observe the correlation between the mesial papilla's height of single implant-supported maxillary central incisor and the distance from the base of the contact point to the alveolar bone crest. METHODS: 56 patients involved in single implant-supported maxillary central incisor were included in this study. The distances from the base of the contact point to the alveolar bone crest in the digital periapical film of maxillary central incisor were measured using the software Planmeca Dimaxis Version 3.3.2. The time of measurements were as follows: The pre-surgical and post-surgical periods, before and after the crown installation, the follow-up examination of more than 0.5 year. To analyze the factor of influencing distance, and the correlation between the distance and the height of gingival papilla during the whole restored period. Correlation analysis between the distance and the height of gingival papilla during the whole restoration was done by the statistical software SPSS 12.0. RESULTS: The results demonstrated that the ratio of esthetic papilla can achieve 54.5% at the crown installation and 95.5% at the follow-up examination when the distance was between 3 mm and 5 mm. When the distance was between 5 mm and 6 mm, they dropped to 30.0% and 75.0%, respectively. However, when the distance increased to above 7 mm, the papilla could hardly be in an esthetic outcome. There was a significant change of the distance was found during the periods from the post-surgery to pre-restoration, and the scope of the changes was between -0.13 mm and 0.46 mm. A negative correlation was found between the distance and the index of papilla. The correlation coefficient r was -0.715 (P < 0.01). CONCLUSION: It is proposed that the pre-surgery distance of maxillary central incisor from the base of the contact point to the alveolar bone crest can be used as one of the important reference indexes to assess and predict the height conditions of gingival papilla.

Comparison of immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) assessment for Her-2 status in breast cancer.

BACKGROUND: The concordance rate between IHC and FISH according to clinical performance is still controversial. We report a prospective study to reflect the concordance between IHC and FISH in Guilin city, People's Republic of China. METHODS: Fifty cases of invasive ductal carcinoma of breast tested by IHC and scored as 0, 1+, 2+ and 3+ by pathologists were further analyzed by FISH using a commercially available double-color probe, and the FISH findings were compared with IHC test results. RESULTS: A total concordance of 82.0% was observed with a Kappa coefficient of 0.640 (P<0.001). A high discordance was observed in 30.0% of the patients with IHC 2+, 7.1% in IHC 3+, 19.2% overall in IHC 0 and 1+. CONCLUSION: The IHC can be used firstly to screen the HER-2 status, and FISH can be used as a supplementary role to IHC and 2+ and some negative cases. And only those cases with Her-2 status of IHC 3+ or FISH positive should be treated with Herceptin.

Dynamic phosphorus budget for lake-watershed ecosystems.

Lake eutrophication caused by excess phosphorus (P) loading from point sources (PS) and nonpoint sources (NPS) is a persistent and serious ecological problem in China. A phosphorus budget, based on material flow analysis (MFA) and system dynamic (SD), is proposed and applied for the agriculture-dominated Qionghai Lake watershed located in southwestern China. The MFA-SD approach will not only cover the transporting process of P in the lake-watershed ecosystems, but also can deal with the changes of P budget due to the dynamics of watershed. P inflows include the fertilizer for agricultural croplands, soil losses, domestic sewage discharges, and the atmospheric disposition such as precipitation and dust sinking. Outflows are consisted of hydrologic export, water resources development, fishery and aquatic plants harvesting. The internal P recycling processes are also considered in this paper. From 1988 to 2015, the total P inflows for Lake Qionghai are in a rapid increase from 35.65 to 78.73 t/a, which results in the rising of P concentration in the lake. Among the total P load 2015, agricultural loss and domestic sewage account for 70.60% and 17.27% respectively, directly related to the rapid social-economic development and the swift urbanization. Future management programs designed to reduce P inputs must be put into practices in the coming years to ensure the ecosystem health in the watershed.