BACKGROUND AND OBJECTIVECombined hypoxic cytotoxic drugs and chemoradiotherapy is an important mean of oncotherapy, and Tirapazamine (TPZ) is one of the most remarkable drugs. It has been shown that TPZ has a synergistic effect with radiotherapy on tumor cells, but whether TPZ would down-regulate the expression of the hypoxia-induced genes has not been reported. This study was to investigate the hypoxia-induced mRNA expressions of hypoxia inducible factor-1alpha (HIF-1alpha) and osteopontin (OPN) in human nasopharyngeal carcinoma HNE-1 and CNE-1 cells and the radiosensitization of TPZ, a hypoxia-specific drug, on HNE-1 and CNE-1 cells in vitro.

METHODSThe IC50 values of TPZ for HNE-1 and CNE-1 cells were measured using MTT assay, and the mRNA expressions of HIF-1alpha and OPN in HNE-1 and CNE-1 cells was determined using RT-PCR under aerobic and hypoxic conditions, respectively. The survival rates of HNE-1 and CNE-1 cells treated with or without TPZ at IC10 in the presence or absence of oxygen for 6 h were determined using colony formation assay following exposure to 1-6 Gy of 60Co radiation. The dose-survival curves were plotted and the values of D0, Dq and SER were calculated as a single-hit multitarget model.

RESULTSThe IC50 values of TPZ were 34.81 μmol/L and 35.02 μmol/L in HNE-1 and CNE-1 cells under aerobic condition, and 30.20 μmol/L and 28.48 μmol/L under hypoxic condition, respectively. The expressions of HIF-1alpha and OPN mRNA were reduced by TPZ in HNE-1 cells, but not in CNE-1 cells under hypoxic condition. For the HNE-1 cells, the respective values of D0 and Dq were 0.89 Gy and 0.28 Gy following normoxic irradiation versus 1.47 Gy and 0.44 Gy following hypoxic irradiation. For the CNE-1 cells, the respective values of D0 and Dq were 0.72 Gy and 0.68 Gy following normoxic irradiation versus 0.95 Gy and 0.56 Gy following hypoxic irradiation. The values of D0 and Dq for HNE-1 and CNE-1 cells treated with TPZ under hypoxic condition following irradiation were 0.66 Gy, 0.21 Gy and 0.85 Gy, 0.79 Gy, respectively.

CONCLUSIONTPZ can down-regulate hypoxia-induced expression of HIF-1alpha and OPN mRNA of HNE-1 cells and radiosensitize the HNE-1 cells but not CNE-1 cells, and act as a hypoxia modifier.

Antineoplastic Agents ; pharmacology ; Cell Hypoxia ; Cell Line, Tumor ; Cell Survival ; drug effects ; radiation effects ; Cobalt Radioisotopes ; Down-Regulation ; Humans ; Hypoxia-Inducible Factor 1, alpha Subunit ; genetics ; metabolism ; Inhibitory Concentration 50 ; Nasopharyngeal Neoplasms ; metabolism ; pathology ; Osteopontin ; genetics ; metabolism ; RNA, Messenger ; metabolism ; Radiation Tolerance ; drug effects ; Radiation-Sensitizing Agents ; pharmacology ; Triazines ; pharmacology

Background and Objective: Combined hypoxic cytotoxic drugs and chemoradiotherapy is an important mean of oncotherapy, and tirapazamine (TPZ) is one of the most remarkable drugs. It has been shown that TPZ has a synergistic effect with radiotherapy on tumor cells, but whether TPZ would down-regulate the expression of the hypoxia-induced genes has not been reported. This study was to investigate the hypoxiainduced expression of hypoxia inducible factor-1α (HIF-1α) and osteopontin (OPN) mRNA in human nasopharyngeal carcinoma HNE-1 and CNE-1 cells and the radiosensitization of TPZ, a hypoxia-specific drug, on HNE-1 and CNE-1 cells in vitro. Methods: The IC_(50) values of TPZ for HNE-1 and CNE-1 cells were measured using MTT assay, and the expression of HIF-1α and OPN mRNA in HNE-1 and CNE-1 cells was determined using RT-PCR under aerobic and hypoxic conditions, respectively. The survival rates of HNE-1 and CNE-1 cells treated with or without TPZ at IC_(10) in the presence or absence of oxygen for 6 h were determined using colony forming assay following exposure to 1-6 Gy of ~(60)Co radiation, and the dose-survival curves were plotted and the values of D_0, D_q and SER were calculated as a singlehit multitarget model. Results: The IC_(50) values of TPZ were 34.81 μmol/L and 35.02 μmol/L in HNE-1 and CNE-1 cells under aerobic condition, and 30.20 μmol/L and 28.48 μmol/L under hypoxic condition. The expressions of HIF-1α and OPN mRNA were reduced by TPZ in HNE-1 cells, but not in CNE-1 cells under hypoxic condition. The values of D_0 and D_q in HNE-1 and CNE-1 cells following irradiation under aerobic and hypoxic conditions were 0.89 Gy, 0.28 Gy and 0.72 Gy, 0.68 Gy, and 1.47 Gy, 0.44 Gy and 0.95 Gy,0.56 Gy, respectively. The values of D_0 and D_q for HNE-1 and CNE-1 cells treated with TPZ under hypoxic condition following irradiation were 0.66 Gy,0.21 Gy and 0.85 Gy, 0.79 Gy, respectively. Conclusion: TPZ would downregulate hypoxia-induced expression of HIF-1α and OPN mRNA of HNE-1 cells and radiosensitize the HNE-1 cells but not CNE-1 cells, and act as a hpoxia modifier.

BACKGROUNDTransplantation of mensenchymal stem cells (MSCs) has been proposed as a promising way for tissue engineering. However, the application of MSCs for transplantation will undergo apoptosis due to the extremely harsh microenvironment such as excessive inflammation. Apigenin (API) has been reported to protect cells against inflammatory damage and cell death by exhibiting anti-inflammatory and anti-oxidative capacity. Here we investigated the modulatory effects of API in lipopolysaccharide (LPS)-mediated inflammation and apoptosis of MSCs, and further defined the underlying mechanism.

METHODSEffects of different concentrations of API (0, 5, 10, 20, 40 and 80 µmol/L) for 24 hours, and LPS (0, 0.5 and 5.0 µg/ml) for 6 hours and 24 hours on MSCs viability were assayed by MTT. Based on this, MSCs were pretreated with different concentrations of API (0 - 40 µmol/L) at the indicated times (6, 12 and 24 hours) followed by exposure to 5 µg/ml LPS for 24 hours. MTT, phase-contrast microscopy, annexinV/propidium iodide (PI) double stain flow cytometry (FCM) and Hoechst staining were applied to explore the effects of API on MSCs induced by 5 µg/ml LPS for 24 hours. In addition, reverse-transcription polymerase chain reaction (RT-PCR) was applied to detect the mRNA expression of pro-inflammatory factors including cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS), nuclear factor-kappa B (NF-κB), pro-apoptotic gene caspase-3, Bad, and anti-apoptotic gene Bcl-2. Moreover, AutoDock software was used to imitate the docking score of API and vitamin D receptor (VDR). In parallel, Western blotting and RT-PCR were used to investigate protein and mRNA expression of VDR.

RESULTSMSCs stimulated with LPS 5 µg/ml for 24 hours was used as a model of apoptosis induced by over inflammatory stimulus. API (0 - 40 µmol/L) had non-toxic effect on MSCs; however, it could decrease mRNA expression of COX-2, iNOS and NF-κB at different time points in MSCs induced by LPS, except for API at the concentration of 5 µmol/L.

RESULTSfrom phase-contrast microscopy, MTT, Hoechst staining and AnnexinV/PI double stain FCM demonstrated that with the increasing concentrations of API and extension of administrating time, significant morphological changes of MSCs occurred, viability of cells was strongly inhibited, and meanwhile, apoptosis of LPS-administrated MSCs was exacerbated, compared with LPS individual group. In addition, API promoted caspase-3, Bad mRNA expression and inhibited Bcl-2 mRNA expression in a time-dependent and concentration- dependent manner. Further study found that pro-apoptosis effect of API was related to suppress VDR expression.

CONCLUSIONSAPI could inhibit the expression of inducible inflammatory factors, therefore exert the strong anti-inflammatory function. However, API could not protect MSC apoptosis induced by LPS but amplified the apoptosis. The apoptosis is related to Bad/Bcl-2 increasing and caspase-3 activation, which is mediated through suppressing VDR expression.

Animals ; Apigenin ; pharmacology ; Apoptosis ; drug effects ; Blotting, Western ; Cell Survival ; drug effects ; Cells, Cultured ; Flow Cytometry ; Lipopolysaccharides ; pharmacology ; Male ; Mesenchymal Stromal Cells ; cytology ; drug effects ; metabolism ; Rats ; Rats, Sprague-Dawley ; Receptors, Calcitriol ; genetics ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction

Objective To investigate the awareness rate,treatment rate and control rate of mineral and bone disorder in patients with moderate or advanced stage chronic kidney disease (CKD). Methods The awareness rate,treatment rate and control rate of mineral and bone disorder were evaluated based on a questionnaire and related laboratory examinations in 503 CKD stage 3 to 5 patients. Results The awareness rate of mineral and bone disorder in patients with moderate or advanced stage CKD was highest in hemodialysis patients,moderate in peritoneal dialysis patients and lowest in non-dialyzed patients (all P ＜0.01).The total scores of the questionnaire were lowest in non-dialyzed patients [6 (5,8)] and were significantly higher in peritoneal dialysis [11 (9,12)] and hemodialysis patients [13 (11,15)] (P＜0.01).The extent of awareness was negatively correlated with age (r=-0.11,P＜0.05),and positively correlated with educational background (r=0.226,P＜0.01),duration of CKD (r=0.597,P＜0.01) and duration of dialysis (r=0.366,P＜0.01).The source of knowledge was mainly from publicity and education made by medical staff,which accounted for 94.0％,79.5％ and 69.4％ respectively in nondialyzed,peritoneal dialysis and hemodialysis patients.The treatment rate was significantly higher in peritoneal dialysis (88.6％) and hemodialysis patients (96.9％) than that in non-dialyzed patients (58.2％) (all P＜0.01).According to K/DOQI guideline,the control rate of serum calcium,phosphorus,calcium and phosphorus product and parathyroid hormone (PTH) were much better in non-dialyzed patients as compared to dialyzed ones.The percentage of number of lab indicators meeting the standard was significantly higher in non-dialyzed patients as compared to dialyzed ones (P＜0.01).According to KDIGO guideline,the control rate of serum phosphorus was significantly lower in hemodialysis patients (23.6％) than that in peritoneal dialysis (36.9％) and non-dialyzed patients (46.7％) (P＜0.01). Conclusions In non-dialyzed patients with moderate or advanced stage CKD,the awareness rate and treatment rate of mineral and bone disorder are relatively low,and the control rate is relatively high.Whereas in dialyzed patients,the awareness rate and treatment rate are relatively high,and the control rate is relatively low.

Objective:To observe heart rate circadian rhythm in patients with chronic kidney disease (CKD) stage 5 and to analyze the effects of parathyroidectomy (PTX) on heart rate circadian rhythm in severe secondary hyperparathyroidism (SHPT) patients.Methods:A cross-sectional observation was performed in 213 patients with CKD stage 5 and 96 controls, and the patients were divided into those with severe SHPT (PTX group, n=70) and without severe SHPT (non-PTX group, n=143). Forty-six PTX patients were followed up prospectively. The baseline data were compared among these groups. Holter electrocardiogram was performed for each participant. Non-dipping heart rate was defined as night/day heart rate ratio greater than 0.9. Multiple linear regression analysis was used to analyze the related factors of heart rate circadian rhythm in patients with CKD stage 5. Results:The 24-hour, daytime and nighttime mean heart rate in patients with CKD stage 5 were all higher than those in controls, especially in PTX group (all P<0.05). The night/day heart rate ratios of controls and CKD stage 5 patients were (0.81±0.08) and (0.91±0.08) respectively ( P<0.01). Correlation analysis showed 24-hour and daytime or nighttime mean heart rate in patients with CKD stage 5 were positively correlated with serum levels of phosphorus and ln(alkaline phosphatase), while nighttime mean heart rate and night/day heart rate ratio were positively related with serum intact parathyroid hormone level. After adjusting with postoperative follow-up period (median time: 10.9 months), 24-hour and nighttime mean heart rate, and night/day heart rate ratio in PTX patients all decreased significantly (all P<0.01). Conclusions:Heart rate is increased and circadian rhythm is abnormal in patients with CKD stage 5, which are related with mineral and bone disorder. PTX significantly decreases 24-hour and nighttime mean heart rate in severe SHPT patients, and improves the heart rate circadian rhythm.

Objective To investigate the specific T cell subpopulation and the relationship with facial motoneuron in immune deficiency mouse model with facial nerve paralysis, so as to find information for new strategy of facial palsy treatment. Methods Firstly, purifying the CD4+ T cell from wild type mouse and reestablishing the immune function of nude mouse by infusing the CD4+ T cell through the tail vein a week before the surgery. Then the all nude mouse(BALB/c background)and wild type mouse(BALB/c background)were subjected to a right facial nerve axotomy. Then the mouse was studied by application and assessment with fluorogold retrotraeer at specific time. After collecting the slices of brain stem three days post the operation,the facial motoneurons was observed under fluorescence microscope, then analyzed and counted with the software Image Pro Plus5.1. Results The number of survival facial motoneuron in the group with CD4+ T cell transplantation and control group was(3444.5±84.2,-x±s)and(3013.2±65.3)respectively. There was significant difference of the number of survival facial motoneurons between nude mouse transplanted with CD4+ T cell and PBS at three days post the operation(t=5.52,P=0.0003).But there was no significant difference of survival facial motoneurons between nude mouse transplanted with CD4+ T cell and wild type mouse three days post the operation(t=0.49,P=0.6347). It was the transplantation of CD4+ T cell that rescued the survival facial motoneuron to the level of wild type.Conclusions CD4+ T cell have the ability to rescue the iniuring facial motoneuron from death. It may suggest that there is a critical role of the specific T cell subpopulation in facial nerve repair and regeneration.

Three bibenzyls 1-3 and six other compounds 4-9 were firstly isolated from Dendrobium huoshanense stems. They were identified as 3',4-dihydroxy-3,5'-dimethoxybibenzyl(1), batatasin Ⅲ(2), 3,4'-dihydroxy-5-methoxy bibenzyl(3), dihydroconiferyl dihydro-p-coumarate(4), syringaresinol(5), 3-(4-hydroxyphenyl)-propionic acid ethyl ester(6),(3-ethylphenyl)-1,2-ethanediol(7),(S)-5-hydroxy-3,4-dimethyl-5-pentylfuran-2(5H)-one(8) and loliolide(9). Anti-inflammation assay showed that bibenzyls 1-3 could significantly inhibit the production of nitric oxide(NO) and the expression of tumor necrosis factor α(TNF-α) and interleukin 1β(IL-1β) mRNA in LPS-induced RAW264.7 macrophages. Mechanism study exhibited that the phosphorylation of nuclear factor kappa B(NF-κB) p65, inhibitor of κB(IκB), extracellular regulatedprotein kinase(ERK), c-Jun N-terminalkinase(JNK), p38 and Akt of LPS-induced RAW264.7 macrophages could be remarkably reduced by 1. These results suggested that the inflammatory response of LPS-induced RAW264.7 macrophages could be significantly inhibited by 1-3. Additionally, the anti-inflammatory effect of 1 might be contributed to its ability on the regulation of NF-κB, MAPKs and Akt signaling pathways.
Anti-Inflammatory Agents ; therapeutic use ; Dendrobium ; Humans ; Inflammation ; drug therapy ; Lipopolysaccharides ; Macrophages ; NF-kappa B ; Nitric Oxide ; Nitric Oxide Synthase Type II

BACKGROUNDAlthough that glomerulonephritis is the major cause of end-stage renal disease in developing countries such as China, the increasing prevalence of diabetes has contributed to the changing spectrum of predialysis chronic kidney disease. Recent studies have revealed an increased proportion of patients with diabetic kidney disease (DKD) in hemodialysis populations in large cities in China. However, studies regarding the clinical phenotype of DKD in China are extremely limited. The incidence, development, and prognosis of diabetic kidney disease (INDEED) study aims to investigate the incidence, progression, and prognosis of DKD, as well as the associated genetic, behavioral, and environmental factors and biomarkers in patients with DKD in China.

METHODSINDEED study is a prospective cohort study based on all participants with diabetes in the Kailuan study, which is a general population-based cohort study in northern China. Altogether, over 10,000 participants with diabetes will be followed biennially. Questionnaires documenting general characteristics, behavioral and environmental factors, and medical history will be administrated. Anthropometric measurements and a series of laboratory tests will be performed in one central laboratory. The DNA, plasma, and urine samples of every participant will be stored in a biobank for future research.

CONCLUSIONSINDEED study will provide essential information regarding the clinical phenotype and prognosis of patients with DKD in China and will be valuable to identify factors and biomarkers associated with patients with DKD in China.

Biomarkers ; China ; epidemiology ; Diabetic Nephropathies ; epidemiology ; pathology ; Female ; Humans ; Incidence ; Male ; Prognosis ; Prospective Studies ; Surveys and Questionnaires

Patients with severe trauma require an extremely timely treatment and transfusion plays an irreplaceable role in the emergency treatment of such patients. An increasing number of evidence-based medicinal evidences and clinical practices suggest that patients with severe traumatic bleeding benefit from early transfusion of low-titer group O whole blood or hemostatic resuscitation with red blood cells, plasma and platelet of a balanced ratio. However, the current domestic mode of blood supply cannot fully meet the requirements of timely and effective blood transfusion for emergency treatment of patients with severe trauma in clinical practice. In order to solve the key problems in blood supply and blood transfusion strategies for emergency treatment of severe trauma, Branch of Clinical Transfusion Medicine of Chinese Medical Association, Group for Trauma Emergency Care and Multiple Injuries of Trauma Branch of Chinese Medical Association, Young Scholar Group of Disaster Medicine Branch of Chinese Medical Association organized domestic experts of blood transfusion medicine and trauma treatment to jointly formulate Chinese expert consensus on blood support mode and blood transfusion strategies for emergency treatment of severe trauma patients ( version 2024). Based on the evidence-based medical evidence and Delphi method of expert consultation and voting, 10 recommendations were put forward from two aspects of blood support mode and transfusion strategies, aiming to provide a reference for transfusion resuscitation in the emergency treatment of severe trauma and further improve the success rate of treatment of patients with severe trauma.