Application of SGRT Combined with IGRT Isocenter Dual-guided Resetting in IMRT for Breast Cancer / 中山大学学报(医学科学版)

ObjectiveThe objective is to investigate the possibility of isocenter dual-guided resetting of surface guided radiation therapy （SGRT） combined with image guided radiation therapy （IGRT） in postoperative radiotherapy for breast cancer. To assess the setup error accuracy between the new resetting mode and the traditional resetting mode. MethodsRetrospective analysis was performed on breast cancer patients who underwent ELEKTA infinity accelerator radiotherapy in sun yat-sen university cancer center from July 13， 2021 to October 15， 2022. According to different reset methods， the patients were divided into a simulation group （41 cases） and a dual-guided group （40 cases）. The simulation group was reset using a simulator， CBCT scans were performed and setup errors were recorded during the first treatment； The dual-guided group was guided by AlignRT and combined with CBCT for isocenter dual-guided resetting， and the setup error obtained by CBCT registration was recorded. The global setup errors of chest region of interest （CROI） ， the local residual errors of supraclavicular region of interest （SROI） and the resetting time of the two reset methods were calculated and compared respectively. The advantages of the CBCT error distribution in the dual-guided resetting of SGRT combined with IGRT were analyzed. ResultsThe median of the global setup errors （X/cm， Y/cm， Z/cm， Rx°， Ry°， Rz°） of the simulation group and the median of the dual-guided group in the CROI were statistically significant （P<0.05） except the Rz and Ry directions. The local residual errors of the two groups of the SROI were calculated. The median of the errors of X/cm， Y/cm， Z/cm， Rx°， Ry°， Rz° were statistically significant （P<0.05） except the X and Y axis. The resetting time of the simulation group was significantly longer than that of the dual-guided group （238.64±28.56） s， t=-24.555， P=0.000， and the difference was statistically significant （P<0.05）. The CBCT error distribution of the dual-guide group was analyzed， and it was found that the absolute values of translation errors of X， Y and Z axis were all within 0.4 cm， while the proportions of ≤ 0.3 cm were 95%， 93% and 93%， respectively. The proportions of rotation errors of Rx， Ry and Rz ≤ 1.5 ° were 90%， 93% and 90%， respectively. ConclusionIn postoperative radiotherapy of breast cancer， SGRT combined with IGRT for isocenter dual-guided resetting can effectively correct the rotational setup errors and residual errors， and improve the accuracy of radiotherapy with less resetting time and high feasibility， which compared with the traditional simulator resetting mode. This precise， unmarked resetting method can be widely used in clinical practice.

Ferroptosis of airway epithelial cells in asthma: current and future / 药学学报

Ferroptosis is a novel cell death mode proposed in recent years, which is characterized by intracellular iron-dependent lipid peroxidation. Its mechanisms include lipid peroxidation, iron accumulation and the imbalance of antioxidant system. The crosstalk between ferroptosis and asthma is gradually deepening. Elucidating the specific mechanism of ferroptosis in regulating asthma is helpful to broaden the understanding of the pathology of asthma. This paper expounds the role of ferroptosis in airway epithelial cells in the occurrence and development of asthma from three perspectives: lipid peroxidation, iron accumulation and the imbalance of antioxidant system, hoping to find new targets and strategies for asthma treatment.

Regulatory mechanism of heat shock protein 90 on autophagy-related transcription factor EB in human hepatocellular carcinoma cells / 生理学报

This study was aimed to investigate the regulatory mechanism of heat shock protein 90 (Hsp90) on transcription factor EB (TFEB) during autophagy in liver cancer cells. Human hepatocellular carcinoma cell line HepG2 was treated with Hsp90 N- and C-terminal inhibitors (STA9090 and Novobiocin), respectively. Western blot and RT-PCR were used to detect the expression levels of TFEB and autophagy-related proteins. Chromatin immunoprecipitation (ChIP) assay was used to observe the ability of Hsp90α binding to the TFEB proximal promoter region. The double-luciferase gene reporter experiment was used to determine the activity of TFEB promoter. The results showed that hypoxia induced up-regulation of TFEB protein and mRNA expression levels in the HepG2 cells. The protein expression levels of TFEB, LC3 and P62 were down-regulated significantly by either STA9090 or Novobiocin, under both normoxic and hypoxic conditions. Transfection of Hsp90α-overexpressing plasmids up-regulated TFEB protein levels in either wild-type or Hsp90α knockout HepG2 cells. Hsp90 bound to the TFEB proximal promoter region and was involved in regulating TFEB transcriptional process. Whereas both STA9090 and Novobiocin inhibited Hsp90 to bind to the TFEB proximal promoter region, and decreased the activity of TFEB promoter. These results suggest that Hsp90 promotes TFEB transcription in human hepatocellular carcinoma cells by binding to the proximal promoter region, thereby up-regulating the expression levels of autophagy-related proteins.

Preliminary study of bone marrow mesenchymal stem cell-derived exo-somes in repair of spinal cord injury in rats / 中国病理生理杂志

AIM:To study the influence of bone marrow mesenchymol stem cell-drived exosomes(BMSC-exo-somes)on hindlimb activity,and the numbers of reactive astrocytes and residual neurons in spinal cord injury(SCI)rats. METHODS:BMSCs were cultured using the whole bone marrow adherent culture method and surface markers CD 90 and CD34 were verified by flow cytometry.Exosomes were isolated by ultracentrifugation and the morphology of exosomes was observed under transmission electron microscope.The protein markers CD63 and CD9 were verified by Western blot.After exosomes were applied to SCI rats,the Basso,Beattie and Bresnahan locomotor rating scale score,the Nissl staining of the lesion site,and the numbers of reactive astrocytes and residual neurons were assessed at various time points.RESULTS:Transmission electron microscopic observation revealed the presence of saucer -shaped vesicles.BMSC-exosomes were found to express high levels of CD63 and CD9.Compared with injury group,significant improvement of hindlimb activity scores from day 14 after injury in treatment group was observed(P<0.05),and less reactive astrocytes and more residual neu-rons from day 7 after injury were also observed(P<0.05).CONCLUSION:BMSC-exosomes inhibit reactive astrocytes and death of neurons,and improve hindlimb activity in the rats after SCI.

Simultaneous determination of 12 chemical components in Tiaojing Pills by RP-HPLC under variable wavelengths / 中草药

Objective: To establish a RP-HPLC method to determine the contents of paeoniflorin, acteoside, ferulic acid, leonurine hydrochloride, hesperidin, paeonol, baicalin, asperosaponin VI, limonin, atractylenolide I, atractylenolide III, and pachymic acid in Tiaojing Pills. Methods: The determination was performed on a Venusil MP-C18 column (250 mm × 4.6 mm, 5.0 μm) with ethanol-acetonitrile (40∶60, A) and 0.2% phosphoric acid (B) as mobile phases for gradient elution, at the flow rate of 0.8 mL/min; The column temperature was 45 ℃. Results: The nine components were well separated and showed good linearity, such as paeoniflorin 0.5-50.0 mg/L (r = 0.999 5), acteoside 0.1-10.0 mg/L (r = 0.999 1), ferulic acid 0.2-20.0 mg/L (r = 0.999 2), leonurine hydrochloride 0.3-30.0 mg/L (r = 0.999 3), hesperidin 4.0-400.0 mg/L (r = 0.999 8), paeonol 0.2-20.0 mg/L (r = 0.999 1), baicalin 0.6-60.0 mg/L (r = 0.999 4), asperosaponin VI 1.5-150.0 mg/L (r = 0.999 8), limonin 7.0-700. 0 mg/L (r = 0.999 9), atractylenolide I 0.5-50. 0 mg/L (r = 0.999 3), atractylenolide III 0.5-50. 0 mg/L (r = 0.999 4), and pachymic acid 1.0-100. 0 mg/L (r = 0.999 6). The precision was good, RSD ≤ 0.97%, the repeatability was good in terms of RSD ≤ 1.25% and the recovery rate was 98.5%-103.5% (RSD ≤ 1.24%). Test solution was stable at room temperature within 24 h. The contents of twelve batches of the paeoniflorin, acteoside, ferulic acid, leonurine hydrochloride, hesperidin, paeonol, baicalin, asperosaponin VI, limonin, atractylenolide I, atractylenolide III and pachymic acid were 4.328-4.688, 0.033-0.054, 0.073-0.091, 0.177-0.199, 0.243-0.283, 0.043-0.069, 1.144-1.173, 0.037-0.061, 0.094-0.126, 0.127-0.157, 0.155-0.179, and 0.285-0.327 mg/g, respectively. Conclusion: The method is rapid and has high sensitivity, high accuracy, and good specificity, It can be applied to the quality control of Tiaojing Pills.

Buyang Huanwu Decoction () reduces infarct volume and enhances estradiol and estradiol receptor concentration in ovariectomized rats after middle cerebral artery occlusion / 中国结合医学杂志

<p><b>OBJECTIVE</b>To investigate the effect of Buyang Huanwu Decoction (, BYHWD) on estradiol (E2) and estradiol receptor (ER) in serum and brain in ovariectomized rats after middle cerebral artery occlusion (MCAO).</p><p><b>METHODS</b>Adult female rats were ovariectomized and focal cerebral ischemic was induced by MCAO. Rats were randomly divided into normal, ovariectomy (OVX), MCAO, OVX+MCAO, OVX+MCAO+E2, and OVX+MCAO+BYHWD group. Rats were administered BYHWD 5 g/kg daily, estradiol valerate 500 μg/kg per day or distilled water for 7 consecutive days. Neuronal function and infarct volume were measured on day 7 after artery occlusion, and E2 and ER concentration in serum and brain were checked by enzyme-linked immunosorbent assay.</p><p><b>RESULTS</b>BYHWD significantly improved the neurological behavior, reduced the infarction volume, increased E2 concentration in serum and brain, and increased ER concentration in the brain in ovariectomized rats after MCAO.</p><p><b>CONCLUSION</b>The neuroprotective effects of BYHWD are associated with estrogen and its receptor.</p>

Classification study on commodity specification and grade standard of Pseudostellariae Radix / 中国中药杂志

<p><b>OBJECTIVE</b>To study the commodity specification and grade standard of Pseudostellaria Radix, for standardizing market order and achieving industrialization, standardization and modernization of Pseudostellariae Radix.</p><p><b>METHOD</b>The different areas and grade medicinal materials of Pseudostellariae Radix were respectively measured in quantitative characteristics of appearance, and the present grade classification of Pseudostellaria Radix was scientifically verified by the content of polysaccharide. Then the determination data were evaluated by spss correlation analysis, principal component analysis and cluster analysis. So combining with the actual production, the commodity grade standard of Pseudostellariae Radix was formulated.</p><p><b>RESULT</b>Correlation analysis indicated that the present grade classification of Pseudostellaria Radix was reasonable, and the more the grade of Pseudostellariae Radix was high, the more the content of polysaccharide was high. Meanwhile, length as a classification index was not suitable for the commodity grade standard of Pseudostellariae Radix. Using principal component analysis and cluster analysis, combining actual production, the thickest diameter, weight of single root tuber and the number of 50 g root tuber were filtrated and the grade was divided into 5 ranks: big, mid- dle, small selected goods, big ungraded goods and small ungraded goods.</p><p><b>CONCLUSION</b>the commodity specification and grade standard of Pseudostellariae Radix that mainly included the thickest diameter, weight of single root tuber and the number of 50 g root tuber was formulated, the standard was divided into 5 grade. Each grade was not only consistent with the present situation of medicinal materials market, it could also reflected the intrinsic quality of Pseudostellariae Radix. In conclusion, the standard could be used as a classifica- tion basis to the commodity specification and grade.</p>

Current study of Toll-like receptor and fundus disease / 中华实验眼科杂志

Toll-like receptors (TLRs) can recognize pathogens associated molecular of micro-organisms,leading to activation of the downstream signal transduction pathway and participate in innate and adaptive immune response.TLRs can also recognize endogenous danger signaling molecule and therefore be involved in pathogenesis of many low-grade chronic inflammatory and autoimmune diseases.TLRs are always expressed on immune cells,and recent studies confirmed that retinal pigment epithelial (RPE) cells also express TLRs.This review focuses on the research progress of fundus diseases and TLRs,such as the recognition of exogenous pathogenic microorganisms and endogenous danger signals in fundus,induction of inflammatory response,phagocytosis of photoreceptor outer segment by RPE and angiogenesis of fundus diseases.

IGF2 expression in blood is not associated with its imprinting status in healthy pregnant Chinese women

Loss of Imprinting (LOI) of IGF2 and over-expressed IGF2 are associated with tumorigenesis. Our previous epidemiological study found a relatively high frequency of IGF2 LOI in healthy mid-gestation pregnant women. The aim of this study is to determine whether the expression of IGF2 is associated with its imprinting status in healthy Chinese pregnant women. The IGF2 imprinting status of 300 pregnant women was analyzed. 20 cases of IGF2 LOI and 20 cases of IGF2 retention of imprinting (ROI) were selected randomly for IGF2 expression analysis. The expression pattern of IGF2 between the group with IGF2 ROI and group with IGF2 LOI in healthy Chinese pregnant women was evaluated by real time PCR and western blot. The result showed no significant differences between IGF2 ROI and LOI groups in mRNA and protein levels. These results imply that IGF2 imprinting status has no obvious impact on its expression. There may be some unknown important factors other than imprinting status driving IGF2 expression.

Therapeutic effect of combined cisplatin and docetaxel vs fluorouracil regimen with concurrent radiotherapy on advanced esophageal carcinoma / 南方医科大学学报

<p><b>OBJECTIVE</b>To compare the therapeutic effect and adverse effects of two regimens, namely cisplatin and docetaxel (DC) regimen and fluorouracil (PF) regimen, both with concurrent radiotherapy, in the treatment of advanced esophageal squamous cancer.</p><p><b>METHODS</b>Forty-eight patients with esophageal squamous cancer were randomly assigned in DC regimen and PF regimen groups. All the patients received conventional radiotherapy at a total dose of 60 Gy (in 30 fractions) for 6 weeks. In DC regimen group, the patients received intravenous infusion of docetaxel (75 mg/m(2)) for 1 h on day 1 and DDP (25 mg/m(2) daily) on days 1-3, with every 28 days as one cycle. PF regimen consisted of cisplatin (25 mg/m(2)) on days 1-3 and continuous intravenous infusion of fluorouracil (500 mg/m(2)) for 5 days, with every 28 days as one cycle. All the patients were suggested to have no less than 2 cycles.</p><p><b>RESULTS</b>The 3-year median survival time in DC regimen was slightly longer than that in PF regimen group (26 vs 23 months, Χ2=3.4041, P=0.065). The same result was also found in the short-term effect and adverse reactions including ?myelosuppression and gastrointestinal reactions. Only the adverse reaction of radiotherapy-induced esophagitis showed a significant difference between the two groups (P=0.049).</p><p><b>CONCLUSION</b>DC regimen with synchronous radiotherapy is effective and safe for treating advanced esophageal squamous cancer.</p>

Combined chemotherapy with cisplatin, docetaxel and capecitabine for metastatic nasopharyngeal carcinoma: a retrospective analysis / 南方医科大学学报

<p><b>OBJECTIVE</b>To evaluate the efficacy and toxicity of the combined chemotherapy with docetaxel, capecitabine and cisplatin (TXP) in the treatment of metastatic nasopharyngeal carcinoma (NPC).</p><p><b>METHODS</b>This retrospective analysis involved 22 patients with metastatic NPC receiving treatment with the TXP regimen. The patients were given docetaxel at 60 mg/m² on day 1, cisplatin at 20 mg/m² on days 1-3, and capecitabine at 1 250 mg/m² on days 1-14, and the treatment cycle was repeated ever 3 weeks.</p><p><b>RESULTS</b>Of the 22 patients, 14 (63%) achieved partial remission, 2 (9%) had complete remission, and 5 (23%) showed stable disease. The overall clinical response rate of the patients was 72% with a 1-year survival rate of 68%, median progression-free survival of 8 months, and overall survival of 14 months. The main toxicity was myelosuppression; 7 (32%) patients experienced grade 3/4 neutropenia, and 5 (23%) had grade 3/4 anemia. All the other adverse effects were tolerable and reversible.</p><p><b>CONCLUSION</b>The TXP regimen is safe and effective for treatment of metastatic NPC, and the results are comparable with those of the reports in recent literatures.</p>

Clinical research of HPV DNA detection and HPV RNA detection in single time / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To research the application of the single time detection of HPV E6/E7 mRNA and HC2-HPV-DNA in cervical screening project.</p><p><b>METHODS</b>We detected both HPV E6/E7 mRNA and HC2-HPV-DNA of each sample which collected from 130 cervical disease patients' cervix during Jan. 2008 and July. 2009. TCT results were taken as standard to evaluate the diagnostic accuracy of the above two test methods in detecting high-grade cervical disease.</p><p><b>RESULTS</b>82.3% (107/130)women were confirmed to infect HPV by HC2-HPV-DNA detection, and 40.0% (52/130) women were confirmed to infect HPV by HPV E6/E7 mRNA detection, there was no significant difference between the two results (chi2 = 24.5, P < 0.05). The sensitivity, specificity, positive predictive value, negative predictive value of HC2-HPV-DNA detection were 90.1%, 22.1%, 37.4% and 82.6%, respectively. The sensitivity, specificity, positive predictive value, negative predictive value of HPV E6/E7 mRNA detection were 65.9%, 73.3%, 55.8% and 80.8%, respectively.</p><p><b>CONCLUSION</b>In clinical cervical screening project of single time, the combination of HC2-HPV-DNA detection and HPV E6/E7 mRNA detection wick take on more potential value than applying each of them alone. RNA;</p>

Construction of underexpression HSP90alpha and overexpression HSP90beta human hepatoma cell line HepG2 / 中国应用生理学杂志

<p><b>AIM</b>To construct underexpression HSP90alpha and overexpression HSP90beta human hepatoma cell line HepG2.</p><p><b>METHODS</b>The combined plasimid pSilencerHSP90alpha and pSmycHSP90beta were introduced into HepG2 by electroporation, respectively. The result of transfection was identified by Western-blotting and the curve of cell growth was drew by MTT. Observe the cell vitality and expression of HSP90.</p><p><b>RESULTS</b>Expression of HSP90 in transfected cell line was shown by Western-blotting: Compared with control, expression of HSP90 in the cells transfected with pSilencerHSP90alpha decreased, whereas that in the cells transfected with pSmycHSP90beta increased.The growth curves of the two groups of transfected cells was as the same as that of the control group.</p><p><b>CONCLUSION</b>The stable overexpression HSP90beta and underexpression HSP90alpha HepG2 cell lines were established.</p>

A review on liquid-filled hard gelatin capsules / 中国中药杂志

Novel dosage forms emerges more and more in recent years. One of them is liquid-filled hard gelatin capsules, which adopt gelatin or the hydroxypropyl methyl cellulose (HPMC) as capsule shell. The liquid-filled hard gelatin capsule is increasingly getting attention because of its new-concept dosage form design, which deliver liquid drugs by solid form. The paper mainly introduces application, pharmaceutical manufacturing, quality assessment, prospect of liquid-filled hard gelatin capsules, and focuses on the application and pharmaceutical manufacturing (preparation) of liquid-filled capsule. It is suggested that the capsule is suitable for various liquid or semi-solid natural plant extract and achieve different release profiles. The preparation adopted liquid-filled hard capsules technology. The influence factors concluded property of shell and device of filling. The quality was often evaluated by moisture content of capsule shell, dissolution rate etc. At the same time, it was pointed out that the new dosage form has remarkable marketing prospect and bring profits for enterprises.

Effect of heat stress and preconditioning on proteomics of NIH-3T3 cells / 南方医科大学学报

<p><b>OBJECTIVE</b>To establish a heat stress adaptation model in mouse fibroblast cell line NIH-3T3, and analyze the effect of stress and adaptation on protein synthesis.</p><p><b>METHODS</b>A heat stress adaptation cell model was established by heat preconditioning at 42 degrees C for 20 min. The total proteins were separated from the cell lysate by two-dimensional electrophoresis (2-DE), and analyzed using PDQUEST software. The effect of heat stress and preconditioning on protein synthesis was studied, and the protein spots related to stress adaptation were identified by peptide mass fingerprinting (PMF).</p><p><b>RESULTS</b>The proteins with increased expressions in cells with heat stress but not prior preconditioning represented mostly proteins with low molecular mass, whereas in cells exposed to heat stress following heat preconditioning, the upregulated proteins showed a wide spectrum of relative molecular mass.</p><p><b>CONCLUSIONS</b>In stress condition, the cells tend to give priority to synthesis of proteins with small molecular mass. Preconditioning of the cells may increase the intracellular reserve of the protective proteins for protection against challenge with potential stress condition.</p>

Expression of glucocorticoid receptor alpha and beta in chronic rhinosinusitis with nasal polyp / 中华耳鼻咽喉头颈外科杂志

<p><b>OBJECTIVE</b>To detect the expression of glucocorticoid receptor (GR) alpha and beta in nasal polyps, and analyze the possible relationship between over-expression of GR beta and steroid insensitivity.</p><p><b>METHODS</b>The expression of GR alpha and GR beta was examined by immunohistochemical SP method in the specimens from 17 patients with recurring nasal polyp, 18 patients with chronic rhinosinusitis and nasal polyp (CRSwNP), and 12 patients with chronic rhinosinusitis without nasal polyp (CRSsNP) that did not recur during follow-up for 1.5 - 2 years.</p><p><b>RESULTS</b>The difference of numbers of GR alpha-positive cells (x +/- s) between groups with recurrent nasal polyp (20.2 +/- 6.9), CRSwNP (20.7 +/-7.2) , CRSsNP (16.9 +/- 7.2) and normal subjects (16.1 +/- 5.3) was not significant (P > 0.05). The numbers of GR beta-positive cells in recurring group (34.2 +/- 7.4) or CRSwNP (31.5 +/- 5.9) were higher than that in CRSsNP (19.8 +/- 7.8) and normal group (10.1 +/- 6.7) respectively (all P < 0.05). There was a trend toward higher level in recurring polyp compared with that of CRSwNP patients without recurrence in follow-up period, although this was not statistically different (P = 0.558). The difference of GR beta/GR alpha ratios (x +/- s) in recurring specimens (1.80 +/- 0.47) and CRSwNP group (1.65 +/- 0.49) was significant compared with normal group (0.77 +/- 0.66) respectively (P < 0.05), while there was no significance compared with CRSsNP (1.23 +/- 0.27, P > 0.05).</p><p><b>CONCLUSIONS</b>The high expression of GR beta in nasal polyp is related to the development of nasal polyp.</p>

Effects of artesunate on CD14 and toll-like receptor 4 in peritoneal macrophages of mice with heat stroke endotoxemia / 中华劳动卫生职业病杂志

<p><b>OBJECTIVE</b>To study the effects of artesunate on CD14 and toll-like receptor 4 (TLR 4) expressions in peritoneal macrophages of mice with heat stroke endotoxemia.</p><p><b>METHODS</b>Kunming mice were randomly divided into the normal temperature group, the hyperthermia group, the normal saline (NS) group and the artesunate group (both i.p.60 mg/kg daily for consecutive five days). The normal temperature group was exposed to the condition of dry bulb temperature (Tdb) 25 degrees C +/- 0.5 degrees C and relative humidity (RH) 43% +/- 5% for 2 hours, while other groups were exposed to the condition of Tdb 35 degrees C +/- 0.5 degrees C and RH 65% +/- 5%. The mRNA expressions of CD14 and TLR 4 in peritoneal macrophages and concentrations of tumor necrosis factor alpha (TNF alpha) in plasma were observed in different time points (1 hour and 2 hour).</p><p><b>RESULTS</b>The mRNA expressions of CD14 and TLR 4 in the normal temperature group were 0.34% +/- 0.047% and 0.31% +/- 0.062% respectively. The expressions of two receptors at 1 hour in the hyperthermia group were significantly increased to 0.53% +/- 0.085% and 0.45% +/- 0.049% compared with the normal group and kept increased at 2 hour (P < 0.01). The mRNA expressions at 1 hour in the NS group were significantly increased but a little bit decreased at 2 hour. The mRNA expressions of CD14 and TLR 4 at 1 hour in the artesunate group were 0.26% +/- 0.051% and 0.25% +/- 0.084% respectively and a little bit decreased at 2 hour. The change of TNF-alpha in each group was almost consistent with the changes of CD14 and TLR 4.</p><p><b>CONCLUSION</b>Artesunate can reduce significantly the expressions of CD14 and TLR 4 in LPS signal transduction pathway and the concentration of TNF-alpha, which perhaps is one of the most important mechanisms that artesunate fights against endotoxemia.</p>

Protective effect of heat preconditioning on NIH-3T3 fibroblast / 中华劳动卫生职业病杂志

<p><b>OBJECTIVE</b>To establish stress adaptation model of mouse fibroblast cell line NIH-3T3, to provide a group of parallel object for stress adaptation research, and to explore the function and mechanism of HSP90 in stress adaptation.</p><p><b>METHODS</b>A stress-adapted cell model was established by thermal preconditioning (42 degrees C, 20 minutes), and the adaptation result was evaluated by observing the change of the membrane injury and the damage of DNA induced by the heat stress for the second time (44 degrees C, 20 minutes). The HSP90 content was detected by Western blot.</p><p><b>RESULTS</b>According to the membrane injury and HSP90 synthesis induced by the heat stress for the second time, it was primarily confirmed that 6 hours after thermal preconditioning were the optimum stress protection time. When cells underwent heat stress for the second time 6 hours after thermal preconditioning, the membrane injury (15.4% +/- 2.6% vs 41.2% +/- 5.1%), damage of DNA (15.1% vs 26.3%) were decreased compared with the control group in which there was no preconditioning. The OD(HSP90)/OD(control) value indicated that the cellular HSP90 contents was decreased immediately after heat stress (44 degrees C, 40 min). The content of HSP90 was 0.82 +/- 0.18 in the heat stress group, 1.70 +/- 0.52 in the preconditioning group and 1.41 +/- 0.16 in the heat stress after preconditioning group.</p><p><b>CONCLUSION</b>With the preconditioning for the NIH-3T3, the time point for the stress protection is confirmed, the model for the cellular stress adaptation is established and the protective effect of HSP90 is primarily confirmed in this model.</p>

Stress response changes of NIH-3T3 cells with HSP90alpha expression inhibition by RNA interference / 南方医科大学学报

<p><b>OBJECTIVE</b>To establish a heat shock protein 90alpha (HSP90alpha) expression-inhibited cell line and study the effect of lowered HSP90alpha level on cell stress response.</p><p><b>METHODS</b>The recombinant plasimid pSilencerHSP90 containing the 21nt small interfering RNA of human HSP90alpha was subcloned, purified and identified by DNA sequence analysis before introduced into mouse fibroblast cell line NIH-3T3 by electroporation. After G418 selection, the positive clones were identified by immunofluorescence and Western blotting. NIH-3T3 cells were subjected to hyperthermia at 44 degrees C for 40 min to simulate oxidative stress, and flow cytometry was performed to analyze the effect of low-level HSP90 on DNA damage under stress condition.</p><p><b>RESULTS</b>Immunofluorescence and Westen blotting showed lowered HSP90 levels in the transfected cells. Compared with the control cells, cells subjected to hyperthermia displayed intensified DNA damage.</p><p><b>CONCLUSION</b>Low-level HSP90alpha causes the cells to be more vulnerable to oxidative stress condition, and HSP90 content can be associated with cell protection against such condition.</p>

Establishment of HSP90 overexpressing cell line and effects on cell proliferation / 中国应用生理学杂志

<p><b>AIM</b>To establish a HSP90 highly expressing cell line and study the effect of high level of HSP90 on cell stress response.</p><p><b>METHODS</b>The recombined plasmid pSmycHSP, which contained the full length DNA coding for human HSP90 B, was introduced into mouse fibroblast cell line NIH-3T3 by electroporation after being subcloned, purified and identified by limited enzyme digestion. After screened by G418, the positive clones were selected and identified by immunocytochemistry and Western-blotting. Contrasted with NIH-3T3 cells transfected with empty plasmid, the effect of high-level HSP90 on cell proliferation and cell cycle was analyzed by MTT method and flow cytometry.</p><p><b>RESULTS</b>The rising level of HSP90 was shown by immunocytochemistry and Western-blotting. Compared with control, the growth of HSP90 highly expressing cell line slowed down and the DNA content of S phase was lower.</p><p><b>CONCLUSION</b>The NIH-3T3 derived cell line, which stably expressed high level of HSP90 was established. The effect of high-level HSP90 on cell proliferation was to retard cell growth by affecting cell cycle.</p>