Influential factors and predictors of anti-N-methyl-D-aspartate receptor encephalitis associated with severity at admission.

OBJECTIVE: We aimed to study the clinical characteristics and biological indicators of anti-N-methyl-D-aspartate receptor (NMDAR) encephalitis with different severity levels to explore factors predicting disease severity at admission. METHODS: Using the modified Rankin scale (mRS), patients were divided into mild-to-moderate group (mRS ≤ 3) and severe group (mRS > 3) on admission based on severity of illness. General information, previous history, premonitory symptoms, clinical manifestations before admission, imaging findings and biochemical tests were compared to explore the clinical manifestations and biological indicators related to the severity of illness at admission. RESULTS: In the severe group, the incidences of fever, anti-infective therapy, generalized seizures, consciousness disorder, blood white blood cell, neutrophils, and neutrophil-lymphocyte ratio (NLR) were higher than those in mild-to-moderate group (P < 0.001, P = 0.001, P = 0.020, P < 0.001, P = 0.002, P < 0.001, P < 0.001, respectively); blood lymphocyte counts was lower than those in mild-to-moderate group (P < 0.001). There was the strongest significant positive correlation between the NLR and disease severity at admission (rs = 0.684, P < 0.001). In multivariate logistic regression, fever, generalized seizures, consciousness disorder, and elevated NLR were independent risk factors for disease severity; the area under the receiver operating characteristic curve was 0.896 (95%CI: 0.840-0.952, P < 0.001). CONCLUSION: Fever, generalized seizures, consciousness disorder, and elevated NLR were independent risk factors for disease severity. NLR is a good predictor of the severity of illness at admission.

Two novel pathogenic variants in the TCOF1 found in two Chinese cases of Treacher Collins syndrome.

BACKGROUND: Treacher Collins Ι syndrome (TCS1, OMIM:154500) is an autosomal dominant disease with a series of clinical manifestations such as craniofacial dysplasia including eye and ear abnormalities, small jaw deformity, cleft lip, as well as repeated respiratory tract infection and conductive hearing loss. Two cases of Treacher Collins syndrome with TCOF1(OMIM:606847) gene variations were reported in the article, with clinical characteristics, gene variants and the etiology. METHODS: The clinical data of two patients with Treacher Collins syndrome caused by TCOF1 gene variation were retrospectively analyzed. The whole exome sequencing (WES) was performed to detect the pathogenic variants of TCOF1 gene in the patients, and the verification of variants were confirmed by Sanger sequencing. RESULTS: Proband 1 presented with bilateral craniofacial deformities, conductive hearing loss and recurrent respiratory tract infection. Proband 2 showed bilateral craniofacial malformations with cleft palate, which harbored similar manifestations in her family. She died soon after birth due to dyspnea and feeding difficulties. WES identified two novel pathogenic variants of TCOF1 gene in two probands, each with one variant. According to the American College of Medical Genetics and Genomics, the heterozygous variation NM_001371623.1: c.877del (p. Ala293Profs*34) of TCOF1 gene was detected in Proband 1, which was evaluated as a likely pathogenic (LP) and de novo variant. Another variant found in Proband 2 was NM_001135243.1: c.1660_1661del (p. D554Qfs*3) heterozygous variation, which was evaluated as a pathogenic variation and the variant inherited from the mother. To date, the two variants have not been reported before. CONCLUSION: Our study found two novel pathogenic variants of TCOF1 gene and clarified the etiology of Treacher Collins syndrome. We also enriched the phenotypic spectrum of Treacher Collins syndrome and TCOF1 gene variation spectrum in the Chinese population, and provided the basis for clinical diagnosis, treatment and genetic counseling.

Cardiotonic Pills® protects from myocardial fibrosis caused by in stent restenosis in miniature pigs.

BACKGROUND: Stent implantation has been increasingly applied for the treatment of obstructive coronary artery disease, which, albeit effective, often harasses patients by in-stent restenosis (ISR). PURPOSE: The present study was to explore the role of compound Chinese medicine Cardiotonic Pills® (CP) in attenuating ISR-evoked myocardial injury and fibrosis. STUDY DESIGN: Chinese miniature pigs were used to establish ISR model by implanting obsolete degradable stents into coronary arteries. Quantitative coronary angiography (QCA) was performed to confirm the success of the model. METHODS: CP was given at 0.2 g/kg daily for 30 days after ISR. On day 30 and 60 after stent implantation, the myocardial infarct and myocardial blood flow (MBF) were assessed. Myocardial histology was evaluated by hematoxylin-eosin and Masson's trichrome staining. The content of ATP, MPO, and the activity of mitochondrial respiratory chain complex Ⅳ were determined by ELISA. Western blot was performed to assess the expression of ATP5D and related signaling proteins, and the mediators of myocardial fibrosis. RESULTS: Treatment with CP diminished myocardial infarct size, retained myocardium structure, attenuated myocardial fibrosis, and restored MBF. CP ameliorated energy metabolism disorder, attenuated TGFß1 up-regulation and reversed its downstream gene expression, such as Smad6 and Smad7, and inhibited the increased expression of MCP-1, PR S19, MMP-2 and MMP-9. CONCLUSION: CP effectively protects myocardial structure and function from ISR challenge, possibly by regulating energy metabolism via inactivation of RhoA/ROCK signaling pathway and inhibition of monocyte chemotaxis and TGF ß1/Smads signaling pathway.

Identification of a TRIM32 from Penaeus monodon is involved in autophagy and innate immunity during white spot syndrome virus infection.

Many tripartite motif (TRIM) family proteins played an important role in regulating innate immune and autophagy pathway and were important for host defenses against viral pathogens. However, the role of TRIM proteins in autophagy and innate immunity during virus infection was seldom studied in crustaceans. In this study, a novel TRIM32 homolog was identified from Penaeus monodon (named PmTRIM32). PmTRIM32 was significantly upregulated by rapamycin stimulation and WSSV infection. RNA interference experiments showed that PmTRIM32 could restrict WSSV replication and lead P. monodon more resistance to WSSV challenge. Autophagy could be induced by WSSV or rapamycin challenge and has been proved to play a positive role in restricting WSSV replication in P. monodon. The autophagy activity induced by WSSV or rapamycin challenge could be obviously inhibited by silence of PmTRIM32 in P. monodon. Further studies revealed that PmTRIM32 positively regulated the expression of nuclear transcription factor (NF-κB) and it mediated antimicrobial peptides. Moreover, Pull-down and in vitro ubiquitination assay demonstrated that PmTRIM32 could interact with WSSV envelope protein and target it for ubiquitination in vitro. Collectively, this study demonstrated that PmTRIM32 restricted WSSV replication and was involved in positively regulating autophagy and NF-κB pathway during WSSV infection in P. monodon.

The Ameliorating Effects of Bushen Huatan Granules and Kunling Wan on Polycystic Ovary Syndrome Induced by Dehydroepiandrosterone in Rats.

AIM: To investigate the effects of Bushen Huatan Granules (BHG) and Kunling Wan (KW), the two Chinese medicines, on the regulation of polycystic ovary syndrome (PCOS) and their underlying mechanisms. MATERIALS AND METHODS: PCOS rat model was established by subcutaneous injection of dehydroepiandrosterone (DHEA) (6 mg/100 g/day) for 20 days, followed by treatment with BHG (0.75, 1.49, and 2.99 g/kg) or KW (0.46, 0.91, and 1.82 g/kg) by gavage for 4 weeks. Estrous cycle was detected by vaginal smears. Follicles development was assessed by histology. Levels of testosterone and insulin in serum were tested by ELISA. Apoptosis of Granulosa cells (GCs) was evaluated by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labeling staining. Pathways associated with apoptosis were detected with western blot. Pregnancy outcome was also assessed. GCs were pre-treated with 10-5 M testosterone in vitro for 24 h, then incubated with serum from rats receiving BHG (1.49 g/kg) or KW (1.82 g/kg). The parameters concerning apoptosis, mitochondrial function and endoplasmic reticulum stress were assessed. RESULTS: Post-treatment with either BHG or KW ameliorated DHEA-induced irregular estrous cycles, follicles development abnormalities, increase of testosterone and insulin in serum, and the apoptosis of GCs. Post-treatment with BHG decreased the expression of cleaved caspase-9/caspase 9, release of cytochrome C from mitochondria, and mitochondria reactive oxygen species production, increased activities of complex I, II, IV of ovarian tissue. Post-treatment with KW decreased the levels of caspase-12, GRP78, C/EBP homologous protein, phosphorylation of IRE-I, x-box-binding protein 1s, as well as phosphorylation of proline-rich receptor-like protein kinase, phosphorylation of eukaryotic translation initiation factor 2α and ATF4 of ovarian tissue and GCs. Both BHG and KW ameliorated pregnancy outcome. CONCLUSION: This study demonstrated BHG or KW as a potential strategy for treatment of PCOS induced by DHEA, and suggested that the beneficial role of the two medicines were mediated by different pathway with the effect of BHG being correlated with the regulation of mitochondria, while the effect of KW being attributable to protection of endoplasmic reticulum stress.

Quantitative Proteomics Reveal That Metabolic Improvement Contributes to the Cardioprotective Effect of T89 on Isoproterenol-Induced Cardiac Injury.

BACKGROUND: T89, a traditional Chinese medicine, has passed phase II, and is undergoing phase III clinical trials for treatment of ischemic cardiovascular disease by the US FDA. However, the role of T89 on isoproterenol (ISO)-induced cardiac injury is unknown. The present study aimed to explore the effect and underlying mechanism of T89 on ISO-induced cardiac injury. METHODS: Male Sprague-Dawley rats received subcutaneous injection of ISO saline solution at 24 h intervals for the first 3 days and then at 48 h intervals for the next 12 days. T89 at dose of 111.6 and 167.4 mg/kg was administrated by gavage for 15 consecutive days. Rat survival rate, cardiac function evaluation, morphological observation, quantitative proteomics, and Western blotting analysis were performed. RESULTS: T89 obviously improved ISO-induced low survival rate, attenuated ISO-evoked cardiac injury, as evidenced by myocardial blood flow, heart function, and morphology. Quantitative proteomics revealed that the cardioprotective effect of T89 relied on the regulation of metabolic pathways, including glycolipid metabolism and energy metabolism. T89 inhibited the enhancement of glycolysis, promoted fatty acid oxidation, and restored mitochondrial oxidative phosphorylation by regulating Eno1, Mcee, Bdh1, Ces1c, Apoc2, Decr1, Acaa2, Cbr4, ND2, Cox 6a, Cox17, ATP5g, and ATP5j, thus alleviated oxidative stress and energy metabolism disorder and ameliorated cardiac injury after ISO. The present study also verified that T89 significantly restrained ISO-induced increase of HSP70/HSP40 and suppressed the phosphorylation of ERK, further restored the expression of CX43, confirming the protective role of T89 in cardiac hypertrophy. Proteomics data are available via ProteomeXchange with identifier PXD024641. CONCLUSION: T89 reduced mortality and improves outcome in the model of ISO-induced cardiac injury and the cardioprotective role of T89 is correlated with the regulation of glycolipid metabolism, recovery of mitochondrial function, and improvement of myocardial energy.

The Composite of 3, 4-Dihydroxyl-Phenyl Lactic Acid and Notoginsenoside R1 Attenuates Myocardial Ischemia and Reperfusion Injury Through Regulating Mitochondrial Respiratory Chain.

AIM: 3,4-Dihydroxyl-phenyl lactic acid (DLA) and notoginsenoside R1 (R1) are known to protect ischemia and reperfusion (I/R) injury by targeting Sirtuin1/NADH dehydrogenase (ubiquinone) 1 alpha subcomplex 10/the Mitochondrial Complex I (Sirt-1/NDUFA10/Complex I) and Rho-associated kinase/adenosine triphosphate (ROCK/ATP) ATP synthase δ subunit (ATP 5D), respectively. We hypothesized that a composite of the two may exhibit a more potent effect on I/R injury. The study was designed to test this hypothesis. MATERIALS AND METHODS: Male Sprague-Dawley rats underwent left anterior descending artery occlusion and reperfusion, with or without DLA, R1, or a combination of 3,4-dihydroxyl-phenyl lactic acid and notoginsenoside R1 (DR) pretreatment. Heart function, myocardial morphology, myocardial infarct, myocardial blood flow (MBF), apoptosis, vascular diameter, and red blood cell (RBC) velocity in venules were evaluated. Myeloperoxidase (MPO), malondialdehyde (MDA), and 8-oxo-deoxyguanosine (8-OHdG) were assessed. The content of ATP, adenosine diphosphate (ADP), and adenosine monophosphate (AMP), the activity of mitochondrial respiratory chain Complex I and its subunit NDUFA10, the Mitochondrial Complex V (Complex V) and its subunit ATP 5D, Sirt-1, Ras homolog gene family, member A (RhoA), ROCK-1, and phosphorylated myosin light chain (P-MLC) were evaluated. R1 binding to Sirt-1 was determined by surface plasmon resonance. RESULTS: DLA inhibited the expression of Sirt-1, the reduction in Complex I activity and its subunit NDUFA10 expression, the increase in MPO, MDA, and 8-OhdG, and apoptosis. R1 inhibited the increase in the expression of RhoA/ROCK-1/P-MLC, the reduction of Complex V activity and its subunit ATP 5D expression, alleviated F-actin, and myocardial fiber rupture. Both DLA and R1 reduced the myocardial infarction size, increased the velocities of RBC in venules, and improved MBF and heart function impaired by I/R. DR exhibited effects similar to what was exerted, respectively, by DLA and R1 in terms of respiratory chain complexes and related signaling and outcomes, and an even more potent effect on myocardial infarct size, RBC velocity, heart function, and MBF than DLA and R1 alone. CONCLUSION: A combination of 3,4-dihydroxyl-phenyl lactic acid and notoginsenoside R1 revealed a more potent effect on I/R injury via the additive effect of DLA and R1, which inhibited not only apoptosis caused by low expression of Sirt-1/NDUFA10/Complex I but also myocardial fiber fracture caused by RhoA/ROCK-1 activation and decreased expression of ATP/ATP 5D/Complex V.

[Chemical Composition Characteristics and Source Apportionment of PM2.5 During the Heating Period of 2016-2017 in the Eastern Part of the North China Plain].

To study the composition characteristics and source apportionment of PM2.5 in the eastern part of the North China Plain, manual samples during the two-year heating period of 2016 and 2017 were collected in seven cities, including Hengshui, Cangzhou, Ji'nan, Dezhou, Binzhou, Zibo, and Liaocheng. The results showed that the average values of ρ(PM2.5) during the observation periods were 137.23 µg·m-3 and 111.83 µg·m-3, respectively, which exceeded the daily average secondary standard limit of GB 3095-2012 "Environmental Air Quality Standard" by 1.8 and 1.5 times, respectively. The mean mass of water-soluble ions accounted for 53.32% and 47.04% of ρ(PM2.5), respectively, of which SNA (NO3-, SO42-, and NH4+) were the main ions. During the 2016 and 2017 observation periods, NO3-/SO42- increased from 1.35 to 1.60, while the concentration of Cl- decreased, indicating a decrease in the impact of coal burning. The proportions of secondary organic carbon (SOC) in organic carbon (OC) were 71.63% and 55.35%, respectively, indicating the source of secondary organic carbon had decreased. Analysis of characteristic elements Fe/Al and Ba/Ni showed that dust sources and vehicle sources had increased significantly. The backward trajectories of air mass results showed that the polluted air mass mainly came from the northwest direction during the observation period. However, the PM2.5 concentration was highest when the air mass came from the Anhui and Jiangsu provinces.

Discovery of cardio-protective constituents of Gualou Xiebai Decoction, a classical traditional Chinese medicinal formula.

BACKGROUD: Finding effective compounds of TCMs has always been the basis for achieving marker-based quality control which is currently most widely used quality control strategy. Gualou Xiebai Decoction (GLXB), a classical TCM formula, is recorded and proven as a therapy for curing coronary heart disease but the effective constituents are unidentified and the substantial basis of the therapeutic effects is not clear. PURPOSE: The present research is an investigation on the chemistry of this formula aiming at finding and precisely identifying effective compounds. STUDY DESIGN AND METHODS: This research started with screening for effective fractions of GLXB by rat myocardial infarction model and H9c2 cell hypoxia/reoxygenation model, then compounds in effective fractions were isolated and identified by phytochemical and spectroscopic methods. The cardio-protective activities of the compounds were tested in vitro and one of the effective compounds was taken as example to investigate the mechanisms. RESULTS: The water-insoluble parts of GLXB were identified as effective parts in both in vitro and in vivo experiments. Systematic isolation of compounds in the effective fractions resulted in the isolation of 34 compounds including 7 new compounds, whereas 8 compounds were effective in protecting H9c2 cells against hypoxia/reoxygenation injury. One of the effective compounds, macrostemonoside P (MP) possibly exerted its effect by activating RISK pathway and attenuating apoptosis. CONCLUSION: An array of effective constituents of GLXB were discovered, and discovery of these compounds contributed to elucidating the substantial basis for the therapeutic effects of this formula, and provides fundaments for establishing Q-markers for further reliable quality control of GLXB.

Post-treatment With Qing-Ying-Tang, a Compound Chinese Medicine Relives Lipopolysaccharide-Induced Cerebral Microcirculation Disturbance in Mice.

Objective: Lipopolysaccharide (LPS) causes microvascular dysfunction, which is a key episode in the pathogenesis of endotoxemia. This work aimed to investigate the effect of Qing-Ying-Tang (QYT), a compound Chinese medicine in cerebral microcirculation disturbance and brain damage induced by LPS. Methods: Male C57/BL6 mice were continuously transfused with LPS (7.5 mg/kg/h) through the left femoral vein for 2 h. QYT (14.3 g/kg) was given orally 2 h after LPS administration. The dynamics of cerebral microcirculation were evaluated by intravital microscopy. Brain tissue edema was assessed by brain water content and Evans Blue leakage. Cytokines in plasma and brain were evaluated by flow cytometry. Confocal microscopy and Western blot were applied to detect the expression of junction and adhesion proteins, and signaling proteins concerned in mouse brain tissue. Results: Post-treatment with QYT significantly ameliorated LPS-induced leukocyte adhesion to microvascular wall and albumin leakage from cerebral venules and brain tissue edema, attenuated the increase of MCP-1, MIP-1α, IL-1α, IL-6, and VCAM-1 in brain tissue and the activation of NF-κB and expression of MMP-9 in brain. QYT ameliorated the downregulation of claudin-5, occludin, JAM-1, ZO-1, collagen IV as well as the expression and phosphorylation of VE-cadherin in mouse brain. Conclusions: This study demonstrated that QYT protected cerebral microvascular barrier from disruption after LPS by acting on the transcellular pathway mediated by caveolae and paracellular pathway mediated by junction proteins. This result suggests QYT as a potential strategy to deal with endotoxemia.

Gualou Xiebai Decoction, a Traditional Chinese Medicine, Prevents Cardiac Reperfusion Injury of Hyperlipidemia Rat via Energy Modulation.

Background: Gualou Xiebai Decoction (GLXB) is a classic prescription of Chinese medicine used for the treatment of cardiac problems. The present study was designed to explore the effect and mechanism of GLXB on ischemia/reperfusion (I/R) induced disorders in myocardial structure and function, focusing on the regulation of energy metabolism and the RhoA/ROCK pathway. Methods: After hyperlipidemic rat model was established by oral administration of high fat diet, the rats were treated with GLXB for 6 weeks and subjected to 30 min occlusion of the left anterior descending coronary artery (LADCA) followed by 90 min reperfusion to elicit I/R challenge. Myocardial infarct size was assessed by Evans blue-TTC staining. Myocardial blood flow (MBF) and cardiac function were evaluated. Enzyme-linked immunosorbent assay was performed to examine the content of ATP, ADP, AMP, CK, CK-MB, LDH, cTnT, cTnI, and IL-6. Double staining of F-actin and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling was conducted to assess myocardial apoptosis. Expressions of ATP synthase subunit δ (ATP 5D), and RhoA and ROCK were determined by Western blotting. Results: Administration with GLXB at high dose for 6 weeks protected heart against I/R-induced MBF decrease, myocardial infarction and apoptosis, ameliorated I/R-caused impairment of cardiac function and myocardial structure, restored the decrease in the ratio of ADP/ATP and AMP/ATP, and the expression of ATP 5D with inhibiting the expression of RhoA and ROCK. Conclusions: Treatment with GLXB effectively protects myocardial structure and function from I/R challenge, possibly via regulating energy metabolism involving inactivation of RhoA/ROCK signaling pathway.

[Anti-tumor constituents from Paris polyphylla var. yunnanensis].

OBJECTIVE: To study the anti-tumor active constituents from the rhizome of Paris polyphylla var. yunnanensis. METHOD: The compounds were isolated by column chromatography with silica gel, and purified by Sephadex LH-20 columu chromafography and reverse-phase preparative, HPLC. The structures were identified by spectroscopic methods. The anti-tumor experiment in vitro, the MTT, was used to screen constituents of P. polyphalla var. yunnanensis. RESULT: Six compounds were obtained and their structures were identified as diosgenin-3-O-alpha-L-arabinofuranosyl (1-->4) -beta-D-glycopyranoside (1), pennogenin-3-O-alpha-L-arabino-furanosyl (1-->4)-beta-D-glycopyranoside (2), isorhamn etin-3-O-beta-D-glycopyranoside (3), ethyl-alpha-D-fructofuranoside (4), pennogenin-3-O-alpha-L-rhamnopyranosyl (1-->4)-[alpha-L-rhamnopyranosyl (1-->2)]-beta-D-glycopyranoside (5) and pennogenin-3-O-alpha-L-rhamnopyranosyl (1-->4)-alpha-L-rhamnopyranosyl (1-->4)-[alpha-L-rhamnopyranosyl (1-->2)]-beta-D-glycopyranoside (6). CONCLUSION: Compounds 1-4 were isolated from this plant for the first time, compounds 3 and 4 was firstly isolated from genus Paris, compound 5 was firstly isolated from the rhizome of this plant. The pharmacological results showed that compounds 1-3, 5 and 6 showed certain inhibition, especially, the activities of compound 5 and 6 were the most significant.

Simultaneous determination of creatine phosphate, creatine and 12 nucleotides in rat heart by LC-MS/MS.

A simple, rapid and sensitive LC-MS/MS method was developed and validated for simultaneous determination of creatine phosphate (CP), creatine (Cr) and 12 nucleotides in rat heart. The analytes, ATP, ADP, AMP, GTP, GDP, GMP, CTP, CDP, CMP, UTP, UDP, UMP, CP, Cr, were extracted from heart tissue with pre-cooled (0°C) methanol/water (1:1, v/v) and separated on a Hypersil Gold AQ C18 column (150mm×4.6mm, 3µm) using an isocratic elution with a mobile phase consisting of 2mmol/L ammonium acetate in water (pH 10.0, adjusted with ammonia). The detection was performed by negative ion electrospray ionization in selective reaction monitoring mode (SRM). In the assay, all the analytes showed good linearity over the investigated concentration range (r>0.99). The accuracy was between 80.7% and 120.6% and the precision expressed in RSD was less than 15.6%. This method was successfully applied to measure the concentrations of the 12 nucleotides, creatine phosphate and creatine in rat heart for the first time.

Two new phenanthrofurans from Pleione bulbocodioides.

Two novel phenanthrofurans, shanciol G (1) and shanciol H (2), together with three known phenolic compounds were isolated from the tubers of Pleione bulbocodioides (Franch) Rolfe. Their structures were elucidated by spectroscopic methods.