Microplastics released from disposable medical devices and their toxic responses in Caenorhabditis elegans.

Owing to accelerated urbanization and industrialization, many plastic products have been manufactured and discharged into the environment, causing environmental and public health problems. Plastics in environmental media are further degraded by prolonged exposure to light, heat, mechanical friction, and other factors to form new pollutants called microplastics (MPs). Medical plastics have become a crucial source of plastics in environmental media. However, the release profiles of MPs from medical plastics and their potential ecological and health risks remain unclear. We used optical photothermal infrared spectroscopy to explore the release profiles of eight typical disposable medical devices under high-temperature steam disinfection (HSD). We also evaluated the toxicity of disposable medical devices-derived MPs in Caenorhabditis elegans (C. elegans). Our results showed that the changes in the surface morphology and modification of the disposable medical devices were mainly associated with the material. Polypropylene (PP) and polystyrene (PS) materials exhibited high aging phenomena (e.g., bumps, depressions, bulges and cracks), and HSD broke their oxygen-containing functional groups and carbon chains. By contrast, minor changes in the chemical and physical properties were observed in the polyvinyl chloride (PVC)-prepared disposable medical devices under the same conditions. Further physicochemical characterization indicated that the amount of MPs released from PP-prepared disposable medical devices (P4: 1.27 ± 0.34 × 106) was greater than that from PVC-prepared disposable medical devices (P7: 1.08 ± 0.14 × 105). The particle size of the released MPs was the opposite, PVC-prepared disposable medical devices (P7: 11.45 ± 1.79 µm) > PP-prepared disposable medical devices (P4: 7.18 ± 0.52 µm). Toxicity assessment revealed that disposable medical devices-released MPs significantly increased germ cell apoptosisin C. elegans. Moreover, MPs from PP-prepared disposable medical devices disrupted the intestinal barrier of worms, decreasing their lifespan. Our findings provided novel information regarding the profiles and mechanisms of MP release from disposable medical devices and revealed their potential risks to ecological environment.

An Antibody Neutralization Determinant on Domain III and the First α-Helical Domain in the Stem-Anchor Region of Tembusu Virus Envelope Protein.

Previous studies identified three neutralizing epitopes on domains I, II, and III of the Tembusu virus (TMUV) envelope (E). More evidence is needed to understand the molecular basis of Ab-mediated neutralization and protection against TMUV. In this study, we observed a neutralizing mAb, 6C8, that neutralized TMUV infection primarily by inhibiting cell attachment. In immunofluorescence assays, 6C8 recognized the premembrane and E proteins coexpressed in HEK-293T cells, but failed to react with premembrane or E expressed individually. Epitope mapping identified nine E protein residues positioned on BC/EF loops and F/G strands in domain III and the first α-helical domain in the stem region. Further investigation with mutant viruses showed that 6C8 pressure resulted in mutations at residues 330 of BC loop and 409 of the first α-helical domain, although 6C8 only exhibited a moderate neutralizing activity in BHK-21 cells and a weak protective activity in BALB/c mice and Shaoxing duck models. Mutations A330S and T409M conferred high- and low-level 6C8 resistance, respectively, whereas the combination of A330S and T409M mutations conferred moderate-level 6C8 resistance. As a result, a quasispecies comprising three groups of antigenic variants appeared in BHK-21 cell-derived viral stocks after repeated passages of TMUV strain Y in the presence of 6C8 treatment. Taken together, these findings have raised a concern about Ab-induced antigenic variations in vivo, and they have revealed information concerning the conformational structure of the 6C8 epitope and its role in constraint on antigenic variations. The present work contributes to a better understanding of the complexity of the TMUV immunogen.

Olfactory Ensheathing Cells Alleviate Facial Pain in Rats with Trigeminal Neuralgia by Inhibiting the Expression of P2X7 Receptor.

Trigeminal neuralgia (TN) is a common facial neuropathic pain that is mainly characterized by spontaneous or induced needling or electric shock pain in the innervation area of the trigeminal nerve. It is also referred to as "the cancer that never dies". The olfactory ensheathing cell (OEC) is a special glial cell in the nervous system that has a strong supportive function in nerve regeneration. Cell transplantation therapy is a useful treatment modality that we believe can be applied in TN management. In this study, OECs were transplanted into the ligation site of the infraorbital nerve of rats. We found that after the OEC transplantation, mechanical pain threshold in the face of the rats was significantly increased. Western blotting, immunofluorescence assay, and reverse transcription-quantitative polymerase chain reaction were performed on the trigeminal ganglia (TG) of model rats. The results revealed a decrease in the expression of P2X7 receptor (P2X7R) in the trigeminal ganglia. Our findings show that OEC transplantation has a good therapeutic effect on TN in rats, and that can reduce the expression of P2X7R in trigeminal ganglia. Therefore, we think that OEC transplantation may be a suitable treatment for TN.

Mapping of a unique epitope on domain III of the envelope protein of Tembusu virus.

We recently developed a Tembusu virus (TMUV)-specific monoclonal antibody (MAb) 12F11, which was found to recognize a long amino acid sequence between residues 8 and 77 of domain III of the envelope protein (EDIII). Here, the epitope recognized by MAb 12F11 was mapped using alanine substitutions combined with dissociation constant analysis. The findings, and prediction of tertiary structure of TMUV EDIII, showed that the MAb 12F11 epitope contained one critical residue and 13 peripheral residues. Moreover, the antigenic site was shown to span four loops (N-terminal region, AB, BC, and CD) and three ß-strands (A, B, and D). The present work contributes to the understanding of antigenic structure of TMUV envelope protein.

Substantial Attenuation of Virulence of Tembusu Virus Strain PS Is Determined by an Arginine at Residue 304 of the Envelope Protein.

The Tembusu virus (TMUV) PS strain, derived by several passages and plaque purifications in BHK-21 cells, displays markedly lower virulence in Pekin ducklings relative to a natural isolate of TMUV, but the potential virulence determinants and the in vivo mechanisms for substantial virulence attenuation of the passage variant remain unknown. Here, we constructed a series of chimeric and mutant viruses and assessed their virulence using a 2-day-old Pekin duckling model. We showed that residue 304 in the envelope (E) protein is the molecular determinant of TMUV virulence. Further investigations with mutant and parental viruses demonstrated that acquisition of positive charges at E protein residue 304 plays a critical role in substantial attenuation of neurovirulence and neuroinvasiveness, which is linked to enhanced binding affinity for glycosaminoglycans (GAGs). In Pekin ducklings infected by subcutaneous inoculation, an Arg at residue 304 in the E protein was shown to contribute to more rapid virus clearance from the circulation, markedly reduced viremia, and significantly decreased viral growth in the extraneural tissues and the central nervous system, relative to a Met at the corresponding residue. These findings suggest that the in vivo mechanism of virulence attenuation of the TMUV passage variant closely resembles that proposed previously for GAG-binding variants of other flaviviruses. Overall, our study provides insight into the molecular basis of TMUV virulence and the in vivo consequences of acquisition of a GAG-binding determinant at residue 304 in the E protein of TMUV.IMPORTANCE TMUV-related disease emerged in 2010 and has a significant economic impact on the duck industry. Although the disease was originally recognized to affect adult ducks, increasing evidence has shown that TMUV also causes severe disease of young ducklings. It is, therefore, essential to investigate the pathogenesis of TMUV infection in a young duckling model. The significance of our studies is in identifying E protein residue Arg304 as the molecular determinant for TMUV virulence and in clarifying the crucial role of positive charges at E protein residue 304 in virulence attenuation of a TMUV passage variant. These data will greatly enhance our understanding of the pathogenesis of TMUV infection in ducklings and have implications for development of a safe and efficient vaccine.

[Therapeutic effect of laparoscopic Roux-en-Y gastric bypass in non-obese patients with type 2 diabetes].

OBJECTIVE: To evaluate the effectiveness of laparoscopic Roux-en-Y gastric bypass (LRYGB) surgery for treatment of type 2 diabetes (T2D) in patients with a body mass index (BMI) < 27.5 kg/m2. METHODS: We retrospectively analyzed the data of patients who underwent LRYGB surgery from March, 2012 to June, 2018 in the General Hospital of Guangzhou Military Command and Jinshazhou Hospital of Guangzhou University of Chinese Medicine. The changes in the parameters of glucose metabolism and physical indicators of the patients in the first, second and third years after the surgery were analyzed in patients in low BMI group and high BMI group. RESULTS: All the 74 patients underwent LRYGB successfully without conversion to open surgery. One year after the surgery, fasting blood glucose (FBG), HbA1c, postprandial blood glucose, fasting insulin, HOMA-IR, fasting C-peptide, BMI, body weight and waistline were significantly improved compared with their preoperative values in low BMI group (P < 0.05). At 2 years after the operation, FBG, HbA1c, postprandial blood glucose, HOMA-IR, BMI, body weight and waistline were significantly improved compared with the preoperative values in low BMI group (P < 0.05). In the third year, FBG, HOMA-IR, fasting C-peptide, body weight and waistline were significantly improved compared with the preoperative values in low BMI group (P < 0.05). There was no significant difference in the parameters of glucose metabolism and islet function between low BMI group and high BMI group at different stages. No serious complications occurred in these patients after the surgery. CONCLUSIONS: LRYGB is effective for treatment of T2D in Chinese patients with a BMI < 27.5. After the surgery, the patient show reduced waistline without significant weight loss. The long-term results of the surgery still require further investigations with a larger samples and longer follow-up.

Single-Anastomosis Duodenal Jejunal Bypass Improve Glucose Metabolism by Regulating Gut Microbiota and Short-Chain Fatty Acids in Goto-Kakisaki Rats.

In recent years, bariatric surgery has emerged as a promising treatment for type 2 diabetes. Bariatric surgery is known to cause alterations in the relative abundance and composition of gut microbiota, which may lead to alterations in the levels of Short-Chain Fatty Acids (SCFAs) that are produced during fermentation by gut microbes. However, little is known about the mechanism of improved glucose metabolism mediated by gut microbiota following bariatric surgery. The aim of our study was to explore whether changes in gut microbiota and in fecal SCFA could be detected following single-anastomosis duodenal jejunal bypass (DJB-sa) surgery, a type of bariatric surgery, and whether these alterations might be related to the improvement of glucose metabolism. To this end, we performed DJB-sa or SHAM surgery on Goto-Kakisaki (GK) rats. We then compared the glucose metabolism as well as changes in gut microbiota and SCFAs levels between both groups. Our results showed that DJB-sa surgery was associated with a significant decrease in fasting blood glucose (FBG), intraperitoneal glucose tolerance test (IPGTT), and fasting serum insulin (FSI). And, DJB-sa led to a change in the composition of gut microbiota including an increase in the relative abundance of SCFA-producing bacteria (Bifidobacterium and Subdoligranulum). Moreover, the levels of six SCFAs in feces, as well as the intestinal expression of SCFA receptors including G-protein-coupled receptor 41 (GPR41), G-protein-coupled receptor 43 (GPR43), and G-protein-coupled receptor 109A (GPR109A), and the expression of Glucagon-like peptide-1 (GLP-1) displayed a significant increase following DJB-sa compared with the Sham group. Thus, the gut microbiota may contribute to the improvement of glucose metabolism in type 2 diabetes following DJB-sa. In conclusion, our study shows that DJB-sa improves glucose metabolism by modulating gut microbiota and by increasing short-chain fatty acid production.

Deleterious Effects of Methylene Blue on Rat Nucleus Pulposus Cell in Vitro: Changes in Cell Viability and Secretory Phenotype in Exposed Cells.

PURPOSE: To investigate the effects of methylene blue (MB) on the viability and secretory phenotype of rat nucleus pulposus (NP) cells in vitro. METHODS: Rat NP cells were isolated, cultured, and treated with different MB concentrations (0-6.25 ng/mL) for different lengths of time. We evaluated the changes in cell morphology and cell viability. We also examined the cells for expression of collagen II, aggrecan, matrix metalloproteinase (MMP)-3, MMP-9, and inducible nitric oxide synthase (iNOS). RESULTS: After 2.5 to 6.25 ng/mL MB induced for 6 hours, numerous NP cells were dyed blue and rounded up. The adherent cell number was reduced by MB treatment. The viability of rat NP cells was significantly inhibited by MB in a dose- and time-dependent manner. Treatment with a very low dose of MB (1.5625 ng/mL) resulted in lower expression of collagen II and aggrecan and higher expression of MMP-3, MMP-9, and iNOS in rat NP cells. CONCLUSIONS: Rat NP cells exposed in vitro to MB significantly reduced their viability. Moreover, MB upregulated catabolism gene expression and downregulated anabolism gene expression in rat NP cells. These results suggest MB may be harmful to NP cells. The dose of intradiskal injected MB should be as low as possible to prevent or limit the damage to intervertebral disks.

Microencapsulated olfactory ensheathing-cell transplantation reduces pain in rats by inhibiting P2X4 receptor overexpression in the dorsal root ganglion.

The aim of this study was to determine the role of microencapsulated olfactory ensheathing-cell transplantation (MC-OEC) in rats in which pain was induced by sciatic nerve injury, and its relationship with the expression level of the P2X4 receptor in the dorsal root ganglion. Olfactory bulb tissues of healthy Sprague-Dawley rats were collected to culture olfactory ensheathing cells using differential attachment methods. Ninety-six healthy Sprague-Dawley rats were randomly assigned to the sham, chronic constriction injury (CCI), olfactory ensheathing cell (OEC), and MC-OEC groups. Mechanical paw withdrawal thresholds were measured 7 and 14 days after surgery. The expression of P2X4 receptor genes in the L4-5 dorsal root ganglion was detected by reverse transcriptase polymerase chain reaction, fluorescence in-situ hybridization, and western blotting. Seven and 14 days after the surgery, the mechanical paw withdrawal thresholds of rats in the MC-OEC, OEC, and CCI groups were decreased compared with the sham group. The expression level of the P2X4 receptor in the L4-5 dorsal root ganglion in CCI, OEC, and MC-OEC groups was increased compared with the sham group. All differences between groups were statistically significant. Transplantation of OEC and MC-OEC can reduce neuropathic pain and inhibit the overexpression of the P2X4 receptor in the L4-5 dorsal root ganglion. The transplantation of MC-OEC was more effective in the MC-OEC group than in the OEC group.

Microencapsulated Schwann cell transplantation inhibits P2X3 receptor expression in dorsal root ganglia and neuropathic pain.

Schwann cell transplantation is a promising method to promote neural repair, and can be used for peripheral nerve protection and myelination. Microcapsule technology largely mitigates immune rejection of transplanted cells. We previously showed that microencapsulated olfactory ensheathing cells can reduce neuropathic pain and we hypothesized that microencapsulated Schwann cells can also inhibit neuropathic pain. Rat Schwann cells were cultured by subculture and then microencapsulated and were tested using a rat chronic constriction injury (CCI) neuropathic pain model. CCI rats were treated with Schwann cells or microencapsulated Schwann cells and were compared with sham and CCI groups. Mechanical withdrawal threshold and thermal withdrawal latency were assessed preoperatively and at 1, 3, 5, 7, 9, 11 and 14 days postoperatively. The expression of P2X3 receptors in L4-5 dorsal root ganglia of the different groups was detected by double-label immunofluorescence on day 14 after surgery. Compared with the chronic constriction injury group, mechanical withdrawal threshold and thermal withdrawal latency were higher, but the expression of P2X3 receptors was remarkably decreased in rats treated with Schwann cells and microencapsulated Schwann cells, especially in the rats transplanted with microencapsulated Schwann cells. The above data show that microencapsulated Schwann cell transplantation inhibits P2X3 receptor expression in L4-5 dorsal root ganglia and neuropathic pain.