This study investigated the effects and underlying mechanisms of vanillic acid(VA) against cardiac fibrosis(CF) induced by isoproterenol(ISO) in mice. Male C57BL/6J mice were randomly divided into control group, VA group(100 mg·kg~(-1), ig), ISO group(10 mg·kg~(-1), sc), ISO + VA group(10 mg·kg~(-1), sc + 100 mg·kg~(-1), ig), ISO + dynamin-related protein 1(Drp1) inhibitor(Mdivi-1) group(10 mg·kg~(-1), sc + 50 mg·kg~(-1), ip), and ISO + VA + Mdivi-1 group(10 mg·kg~(-1), sc + 100 mg·kg~(-1), ig + 50 mg·kg~(-1), ip). The treatment groups received the corresponding medications once daily for 14 consecutive days. On the day after the last administration, cardiac functions were evaluated, and serum and cardiac tissue samples were collected. These samples were analyzed for serum aspartate aminotransferase(AST), lactate dehydrogenase(LDH), creatine kinase-MB(CK-MB), cardiac troponin I(cTnI), reactive oxygen species(ROS), interleukin(IL)-1β, IL-4, IL-6, IL-10, IL-18, and tumor necrosis factor-α(TNF-α) levels, as well as cardiac tissue catalase(CAT), glutathione(GSH), malondialdehyde(MDA), myeloperoxidase(MPO), superoxide dismutase(SOD), total antioxidant capacity(T-AOC) activities, and cytochrome C levels in mitochondria and cytoplasm. Hematoxylin-eosin, Masson, uranium acetate and lead citrate staining were used to observe morphological and mitochondrial ultrastructural changes in the cardiac tissues, and myocardial injury area and collagen volume fraction were calculated. Flow cytometry was applied to detect the relative content and M1/M2 polarization of cardiac macrophages. The mRNA expression levels of macrophage polarization markers [CD86, CD206, arginase 1(Arg-1), inducible nitric oxide synthase(iNOS)], CF markers [type Ⅰ collagen(Coll Ⅰ), Coll Ⅲ, α-smooth muscle actin(α-SMA)], and cytokines(IL-1β, IL-4, IL-6, IL-10, IL-18, TNF-α) in cardiac tissues were determined by quantitative real-time PCR. Western blot was used to detect the protein expression levels of Coll Ⅰ, Coll Ⅲ, α-SMA, Drp1, p-Drp1, voltage-dependent anion channel(VDAC), hexokinase 1(HK1), NOD-like receptor protein 3(NLRP3), apoptosis-associated speck-like protein(ASC), caspase-1, cleaved-caspase-1, gasdermin D(GSDMD), cleaved N-terminal gasdermin D(GSDMD-N), IL-1β, IL-18, B-cell lymphoma-2(Bcl-2), B-cell lymphoma-xl(Bcl-xl), Bcl-2-associated death promoter(Bad), Bcl-2-associated X protein(Bax), apoptotic protease activating factor-1(Apaf-1), pro-caspase-3, cleaved-caspase-3, pro-caspase-9, cleaved-caspase-9, poly(ADP-ribose) polymerase-1(PARP-1), and cleaved-PARP-1 in cardiac tissues. The results showed that VA significantly improved cardiac function in mice with CF, reduced myocardial injury area and cardiac index, and decreased serum levels of AST, CK-MB, cTnI, LDH, ROS, IL-1β, IL-6, IL-18, and TNF-α. VA also lowered MDA and MPO levels, mRNA expressions of IL-1β, IL-6, IL-18, and TNF-α, and mRNA and protein expressions of Coll Ⅰ, Coll Ⅲ, and α-SMA in cardiac tissues, and increased serum levels of IL-4 and IL-10, cardiac tissue levels of CAT, GSH, SOD, and T-AOC, and mRNA expressions of IL-4 and IL-10. Additionally, VA ameliorated cardiac pathological damage, inhibited myocardial cell apoptosis, inflammatory infiltration, and collagen fiber deposition, reduced collagen volume fraction, and alleviated mitochondrial damage. VA decreased the ratio of F4/80~+CD86~+ M1 cells and the mRNA expressions of CD86 and iNOS in cardiac tissue, and increased the ratio of F4/80~+CD206~+ M2 cells and the mRNA expressions of CD206 and Arg-1. VA also reduced protein expressions of p-Drp1, VDAC, NLRP3, ASC, caspase-1, cleaved-caspase-1, GSDMD, GSDMD-N, IL-1β, IL-18, Bad, Bax, Apaf-1, cleaved-caspase-3, cleaved-caspase-9, cleaved-PARP-1, and cytoplasmic cytochrome C, and increased the expressions of HK1, Bcl-2, Bcl-xl, pro-caspase-3, pro-caspase-9 proteins, as well as the Bcl-2/Bax and Bcl-xl/Bad ratios and mitochondrial cytochrome C content. These results indicate that VA has a significant ameliorative effect on ISO-induced CF in mice, alleviates ISO-induced oxidative damage and inflammatory response, and its mechanism may be closely related to the inhibition of Drp1/HK1/NLRP3 and mitochondrial apoptosis signaling pathways, suppression of myocardial cell inflammatory infiltration and collagen fiber deposition, reduction of collagen volume fraction and CollⅠ, Coll Ⅲ, and α-SMA expressions, thus mitigating CF.
Animals ; Isoproterenol/adverse effects* ; Male ; Mice ; Signal Transduction/drug effects* ; Vanillic Acid/administration & dosage* ; Dynamins/genetics* ; Mice, Inbred C57BL ; Fibrosis/genetics* ; Apoptosis/drug effects* ; Mitochondria/metabolism* ; NLR Family, Pyrin Domain-Containing 3 Protein/genetics* ; Myocardium/metabolism* ; Humans

As anti-aging ingredients, β-nicotinamide mononucleotide(NMN) and nicotinamide adenine dinucleotide(NAD~+) have attracted worldwide attention in recent years. After oral administration, NMN can be converted into NAD~+ in vivo and the latter is the actual ingredient which exerts anti-aging effect. In order to explore the "rejuvenating and anti-aging" effect of Dendrobium officinale, which was firstly recorded in Shennong's Herbal Classic of Materia Medica, this study established the quantitative method of UPLC-MS/MS for simultaneous determination of NMN and NAD~+ in D. officinale and the congeneric species for the first time, and 34 batches of samples were detected. UPLC conditions are as follows: ACQUITY UPLC HSS T3 column(2.1 mm × 100 mm, 1.8 μm), gradient elution with acetonitrile-0.1% formic acid in water at the flow rate of 0.3 mL·min~(-1), and column temperature of 40 ℃. MS conditions were scanned electrospray ionization source and multiple reaction monitoring mode. The method was verified by systematic methodology. The mean recoveries of NMN and NAD~+ were 77.58% and 80.70%, respectively, with RSD of 3.6% and 4.3%, separately. All results showed that the content of NMN was higher in D. officinale than in the other congeneric species. Particularly, the content in fresh D. officinale stems was as high as 0.931 9 μg·g~(-1). NAD~+ was only found in D. officinale and the content was three times higher than that of NMN. This may be the reason that D. officinale topped the "nine famous anti-aging herbs". In addition, processing method influences the content of NMN and NAD~+ in Dendrobium. Specifically, the content of NMN and NAD~+ was in the order of fresh Dendrobium stems > dried Dendrobium stem segments > spiral or spring-like dried Dendrobium stems.
Chromatography, High Pressure Liquid ; Chromatography, Liquid ; Dendrobium ; NAD ; Nicotinamide Mononucleotide ; Tandem Mass Spectrometry

Taking Lonicerae Japonicae Flos as an example, the method of "expert consensus of different regions" was used to screen the representative samples and evaluate their commodity grades. The correlation analysis, hierarchical cluster analysis and partial least squares discriminant analysis(PLS-DA) of "commodity grade-appearance characteristic-component content" were carried out to reveal the scientificity of traditional commodity grade of Chinese medicinal material. By referring to the existing literature and the grade investigation from the sample collection regions, 78 "initial grade" samples were screened out from 118 collected samples. Authoritative experts from four regions(n=4) including Linyi(Shangdong province), Bozhou(Anhui province), Anguo(Hebei province) and Beijing were organized to evaluate their commodity grades, separately. Based on the grade consistency rate(R_i≥70%), 69 "local grade" samples were screened out from the "initial grade" samples. Based on the average grade consistency rate ■ "authoritative grade" samples were screened out from the "local grade" samples, including15 first-grade samples, 9 second-grade samples, 11 third-grade samples and 17 fourth-grade samples. For these "authoritative grade" samples, the main appea-rance characteristics were quantified and the contents of 13 components were determined by ultra performance liquid chromatography(UPLC). Furthermore, the total contents of 6 phenolic acids, 4 flavonoids and 3 iridoids were calculated, respectively. The results of correlation analysis showed that 4 appearance characteristics indices were correlated with the commodity grades: color, rate of yellow bars(including blooming flowers), rate of black heads(including black bars), and rate of stems and leaves(including bud debris). Five component content indices were correlated with the commodity grades: chlorogenic acid, isochlorogenic acid C, sweroside, loganin and the total contents of six phenolic acids. Furthermore, chlorogenic acid, loganin and the total contents of six phenolic acids showed significantly negative correlation with the main appearance characteristics, indicating that the appearance characteristics of Lonicerae Japonicae Flos can reflect its internal quality, and these 3 indices can be used as quality markers(Q-markers). The results of hierarchical cluster analysis showed that the samples of four grades were classified into four categories, and the samples with the same grades and the same categories accounted for 80.8% of the total samples, while the samples with the different grades were obviously classified into different categories. The results of PLS-DA analysis showed that the samples of different grades showed obvious intragroup aggregation and intergroup dispersion. The above results indicated that it was feasible to evaluate the traditional commodity grade of Lonicerae Japonicae Flos by the method of "expert consensus of different regions". For the evaluation of traditional commodity grade of Chinese medicinal material, the samples should be representative, expert conclusions should have enough consensuses, and grade determination should be authoritative. As the crystallization of clinical experience, traditional commodity grade can scientifically reflect the internal quality of Chinese medicinal material.
Chromatography, High Pressure Liquid ; Drugs, Chinese Herbal ; Flowers ; Lonicera ; Quality Control

Objective To evaluate the relationship between mechanical ventilation-induced apoptosis in hippocampal neurons and mammalian taget of rapamycin ( mTOR) signaling pathway in mice. Methods Fifty healthy male C57BL∕6 mice, aged 8-10 weeks, weighing 20-25 g, were divided into 2 groups ( n=25 each) using a random number table method: control group ( group C ) and mechanical ventilation group ( group V) . The mice breathed spontaneously for 6 h in group C, and the mice were mechanically ventilated for 6 h in group V. Open field test and contextual fear conditioning test were conducted at 1 and 3 days after the end of ventilation. Hippocampal tissues were obtained at 1 day after the end of ventilation for determina-tion of the expression of mTOR, phosphorylated mTOR (p-mTOR), microtubule-associated protein 1 light chain 3Ⅱ( by Western blot) and apoptosis in hippocampal neurons ( by TUNEL) . The p-mTOR∕mTOR ratio and apoptosis index were calculated. Results Compared with group C, the time animals spent in the central square was significantly prolonged, the number of crossing the grid was reduced, the percentage of freezing time was decreased, the expression of microtubule-associated protein 1 light chain 3Ⅱwas up-regulated, and the p-mTOR∕mTOR ratio and apoptosis index were increased in group V ( P<0. 05) . Conclusion The mech-anism by which mechanical ventilation induces apoptosis in hippocampal neurons may be related to activation of mTOR signaling pathway in mice.

Objective: To explore the effect of Feng-liao-chang-wei-kang (FLCWK) on acute and chronic gastroenteritis, synergistic effect on the growth inhibitory effect with 5-fluorouracil (5-FU) on colorectal cancer and its underlying mechanisms. Methods: In the in vitro study, HT-29 cells were divided into 5-FU alone, FLCWK alone and coadministration groups. The MTT assay was used to analyze the proliferation of HT-29 cells at 24 h. Flow cytometry was used to observe the apoptosis, cycle of colorectal cancer HT-29 cells at 24 h. In the in vivo experiment, The subcutaneous transplantation tumor model of colorectal cancer HT-29-Luc was established with nude mice. All mice were randomly divided into 5-FU alone, FLCWK alone and coadministration groups according to body weight. During administration, the Interactive Video Information System small animal live imaging system was used to monitor the growth of subcutaneous transplantation in nude mice. The model of colitis-associated colorectal cancer (CACC) was established with BALB/c mice. BALB/c mice were randomly divided into the normal control group, the model control group, and the FLCWK group. At the end of the administration, the pathological status was detected by HE staining. Cell apoptosis of tumor tissue tumor and colon tissues were observed by TUNEL staining and TUNEL green fluorescence. The protein expression of Caspase 3, p-STAT3, Bcl-2, Bax and P-gp in tumor tissues tumor and colon tissues were tested by using immunohistochemical assay. Results: FLCWK and 5-FU coadministration suppressed HT-29 cell viability and induced S phase arrest and apoptosis compared to treatment with 5-FU alone. Furthermore, compared to treatment with 5-FU alone, coadministration of FLCWK and 5-FU obviously reduced tumor volume and weight and induced apoptosis through decreasing p-STAT3 and P-gp and increasing Caspase 3 protein expression in a murine xenograft tumor model. Moreover, the result revealed decreased number and size of tumors following FLCWK protective administration, downregulated p-STAT3 and Bcl-2 levels and upregulated Bax and Caspase 3 expression in mice with CACC. Conclusions: FLCWK has synergistic effects with 5-FU on colorectal cancer by suppressing the STAT3 pathway and downregulating P-gp expression. Furthermore, FLCWK administration suppresses CACC tumorigenesis by inhibiting the STAT3 pathway.

OBJECTIVETo analyze the clinical manifestations, imaging features and pathological diagnosis of patients with primary central nervous system lymphoma.

METHODSThe clinical data of 50 patients with primary central nervous system lymphoma admitted in our hospital from February 2016 to February 2008 were retrospectively analyzed. All the patients were examined by routine pathology and immunohistochemical staining. Among them 15 cases were examined by MVD and VEGF, and the other 15 glioma patients were taken as control group.

RESULTSIn 50 patients, the disease was chronic, and the main clinical symptoms were numbness, cognitive disorder and disorder of consciousnessetc. Brain CT image of 33 cases (66%) mainly showed slightly higher density; 46 cases (92%) had head enhanced MRI lesions; 38 cases (76%) showed intracranial multiple lesions, 36 cases (72%) showed invasion of supratentorial, and 11 cases showed midline invasion (22%). Pathological diagnosis confirmed 47 cases (94%) with diffuse large B cell lymphoma, the proliferation index of the Ki-67(90%) in 41 case (82%) was higher.

CONCLUSIONPrimary central nervous system lymphoma is manifested with diffuse large B cell lymphoma as its main type, or with complicated clinical manifestations, lacks of features and certain imaging characteristics, but a few patients are easily pathologically misdiagnosed, therefore the biopsy is necessary for diagnosis of these patients.

PURPOSE: Previous studies reported an association between an increased risk of tongue cancer and radiation treatment for nasopharyngeal carcinoma (NPC). This study compared the clinicopathologic characteristics and outcomes of tongue squamous cell carcinoma (TSCC) in patients with and without a history of radiotherapy for NPC. MATERIALS AND METHODS: From 1965 to 2009, a total of 73 patients were diagnosed with TSCC with a history of radiotherapy for NPC. The patients were matched in a 1:3 ratio with patients with sporadic TSCC according to age, sex, and year of the TSCC diagnosis. The primary endpoint was the overall survival. RESULTS: The median interval from NPC to TSCC was 82 months. The NPC survivors were more likely to be diagnosed with a more advanced T classification, less likely to have lymph node involvement, and more likely to have the tumor located in the dorsum of the tongue than sporadic TSCC. Regarding the histologic characteristics, the NPC survivors were more likely to have a weak lymphocytic host response, low tumor budding, and low risk of a worse pattern of invasion. The sporadic TSCC patients had a better overall survival (hazard ratio, 0.690; p=0.033) than the NPC survivors. In competing risks analysis, the cumulative incidence functions for the competing event (documented non-tongue cancer death) were significantly higher in the NPC survivors (Gray's test, p=0.001). CONCLUSION: TSCC patients with a history of radiotherapy for NPC appear to have particular clinicopathologic features, a poorer survival, and are more likely to die from non-tongue cancer causes than those with sporadic TSCC.
Carcinoma, Squamous Cell* ; Case-Control Studies* ; Classification ; Diagnosis ; Epithelial Cells* ; Humans ; Incidence ; Lymph Nodes ; Neoplasms, Second Primary ; Prognosis* ; Radiotherapy ; Survivors ; Tongue Neoplasms ; Tongue*

INTRODUCTIONThis study aimed to report the injury or disease patterns, challenges, key observations, and recommendations by the Singapore Armed Forces (SAF) team that embarked on an Humanitarian Assistance and Disaster Relief (HADR) mission in the aftermath of the April 2015 Nepal earthquake.

METHODSThe SAF medical team that provided HADR assistance to Nepal consisted of personnel from the SAF, Singapore¢s Ministry of Health and the Royal Brunei Armed Forces. Upon arrival in Kathmandu, Nepal, the SAF medical team was assigned to the Gokarna district by the local health authorities. In addition to providing primary healthcare, the medical facility was equipped to perform resuscitation and minor procedures. We also assembled mobile medical teams (MMTs) that travelled to various remote areas of the country to deliver medical aid.

RESULTSA total of 3,014 patients were managed by the SAF medical team. Of these patients, 1,286 (42.7%) were men. 574 (19.0%) patients sustained earthquake-related injuries or illnesses, while 2,440 (81.0%) sustained non-earthquake-related injuries or illnesses. The team treated a total of 447 (77.9%) adults and 127 (22.1%) paediatric patients with earthquake-related injuries or illnesses. A significant number of patients developed exacerbations of underlying medical conditions. 2,161 (71.7%) patients were treated in our main facility in Gokarna, while 853 patients (28.3%) were treated by our MMTs.

CONCLUSIONThe ability to transport healthcare personnel and essential medical equipment within a short time allowed the SAF medical team to provide crucial medical care in the aftermath of the 2015 Nepal earthquake.

Adolescent ; Adult ; Aged ; Brunei ; Child ; Child, Preschool ; Disasters ; Earthquakes ; Emergency Medicine ; Female ; Humans ; Infant ; Infant, Newborn ; Male ; Middle Aged ; Military Personnel ; Nepal ; Physical Examination ; Primary Health Care ; Relief Work ; Singapore

BACKGROUNDInsulin-like growth factor-binding protein-3 (IGFBP-3) is suggested to predict the radiosensitivity and/or prognosis of patients with esophageal squamous cell carcinoma (ESCC). The present study was designed to investigate the clinical and prognostic effects of IGFBP-3 on ESCC.

METHODSIGFBP-3 was detected by immunohistochemistry in paraffin-embedded tissues from 70 ESCC patients treated with radiotherapy alone and further examined by western blotting analysis in 10 pairs of fresh ESCC tissues and adjacent non-malignant esophageal specimens. Receiver operating characteristic (ROC) analysis was used to determine cut-off scores for tumor positivity and to evaluate patient survival status. The χ(2) test was performed to analyze the association of IGFBP-3 expression with clinical characteristics and radiotherapy response. Associations between prognostic outcomes and IGFBP-3 expression were investigated using Kaplan-Meier analysis and the Cox proportional hazards model.

RESULTSThe threshold for IGFBP-3 positivity was set to greater than 65% [area under the ROC curve (AUC)=0.690, P<0.019]. Of the 70 ESCC patient tissues tested, 32 (45.7%) were defined as having high IGFBP-3 expression. The levels of IGFBP-3 protein expression were decreased in 70.0% (7 of 10) of ESCC tissues compared with adjacent non-malignant esophageal tissue. In addition, IGFBP-3 expression was associated with pathologic classification (P<0.05 for T, N, and M categories and clinical stage). Patients with elevated protein level of IGFBP-3 in the tumor had an improved radiotherapy response and prolonged overall survival (P<0.001).

CONCLUSIONSHigh level of IGFBP-3 expression in ESCC associates with early clinical stages and are predictive for favorable survival of the patients treated with radiotherapy.

Blotting, Western ; Carcinoma, Squamous Cell ; Esophageal Neoplasms ; Humans ; Immunohistochemistry ; Insulin-Like Growth Factor Binding Protein 3 ; Kaplan-Meier Estimate ; Prognosis ; Proportional Hazards Models ; ROC Curve ; Radiation-Sensitizing Agents

Objective To evaluate the role of endothelial nitric oxide synthase (eNOS) in inflammatory responses in mice with ventilator-induced lung injury using gene knockout.Methods Twenty-four male C578L/6J wild type mice and 24 male B6.129P2-Nos3tm1Unc/NJU (eNOS gene knockout) mice,aged 10-12 months,weighing 20-25 g,were randomly assigned into 2 groups (n=12 each) using a random number table:sham operation group (group S) and mechanical ventilation group (group MV).The animals were anesthetized with intraperitoneal 1％ pentobarbital sodium 70 mg/kg,and mechanically ventilated after tracheal intubation.The animals were mechanically ventilated for 4 h (oxygen flow rate 0.5 L/min,FiO2 50％,VT 15 ml/kg,RR 70 bpm,PEEP 2 cmH2O).After 4 h of ventilation,blood samples were obtained from the internal carotid artery for detection of PaO2.The animals were then sacrificed and the lungs were removed for determination of wet/dry lung weight (W/D) ratio,myeloperoxidase (MPO) activity,contents of malondialdehyde (MDA),interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α),and pulmonary microvascular permeability,and for microscopic examination of the pathological changes of lung (with electron microscope).Results Compared with group S of wild type mice,PaO2 was significantly decreased,while W/D ratio,MPO activity,contents of MDA,IL-6,TNF-oα and NO,and pulmonary microvascular permeability were increased in MV groups of wild type and gene knockout mice,and no significant change was found in the parameters mentioned above in group S of gene knockout mice.Compared with group MV of wild type mice,PaO2 was significantly increased,while W/D ratio,MPO activity,contents of MDA,IL-6,TNF-α and NO and pulmonary microvascular permeability were decreased in group MV of gene knockout mice.The pathological changes of lung were significantly attenuated in group MV of gene knockout mice as compared with group MV of wild type mice.Conclusion eNOS is involved in inflammatory responses in mice with ventilator-induced lung injury.