C-fos is a transcription factor that plays an important role in cell proliferation, differentiation and tumor formation. The aim of this study was to clone the goat c-fos gene, clarify its biological characteristics, and further reveal its regulatory role in the differentiation of goat subcutaneous adipocytes. We cloned the c-fos gene from subcutaneous adipose tissue of Jianzhou big-eared goats by reverse transcription-polymerase chain reaction (RT-PCR) and analyzed its biological characteristics. Using real-time quantitative PCR (qPCR), we detected the expression of c-fos gene in the heart, liver, spleen, lung, kidney, subcutaneous fat, longissimus dorsi and subcutaneous adipocytes of goat upon induced differentiation for 0 h to 120 h. The goat overexpression vector pEGFP-c-fos was constructed and transfected into the subcutaneous preadipocytes for induced differentiation. The morphological changes of lipid droplet accumulation were observed by oil red O staining and bodipy staining. Furthermore, qPCR was used to test the relative mRNA level of the c-fos overexpression on adipogenic differentiation marker genes. The results showed that the cloned goat c-fos gene was 1 477 bp in length, in which the coding sequence was 1 143 bp, encoding a protein of 380 amino acids. Protein structure analysis showed that goat FOS protein has a basic leucine zipper structure, and subcellular localization prediction suggested that it was mainly distributed in the nucleus. The relative expression level of c-fos was higher in the subcutaneous adipose tissue of goats (P < 0.05), and the expression level of c-fos was significantly increased upon induced differentiation of subcutaneous preadipocyte for 48 h (P < 0.01). Overexpression of c-fos significantly inhibited the lipid droplets formation in goat subcutaneous adipocytes, significantly decreased the relative expression levels of the AP2 and C/EBPβ lipogenic marker genes (P < 0.01). Moreover, AP2 and C/EBPβ promoter are predicted to have multiple binding sites. In conclusion, the results indicated that c-fos gene was a negative regulatory factor of subcutaneous adipocyte differentiation in goats, and it might regulate the expression of AP2 and C/EBPβ gene expression.
Animals ; Goats/genetics* ; Cell Differentiation/genetics* ; Adipogenesis/genetics* ; Gene Expression Regulation ; Proteins/genetics* ; Cloning, Molecular

Objective:To analyze the relationship between gene polymorphisms of methylenetetrahydrofolate reductase (MTHFR) and plasminogen activator inhibitor-1 (PAI-1) and lower limb deep venous thrombosis (DVT) after colorectal cancer surgery.Methods:The clinical data of patients with colorectal cancer who underwent surgical treatment in Zhangjiakou First Hospital from February 2020 to February 2023 were analyzed retrospectively. According to the presence or absence of lower limb DVT after surgery, the patients were divided into DVT group (20 cases) and non-DVT group (80 cases). The polymorphism of MTHFR C677T and PAI-1 promoter 4G/5G were detected by polymerase chain reaction method. The relationship between the polymorphisms of MTHFR C677T and PAI-1 promoter 4G/5G and lower limb DVT after colorectal cancer surgery was discussed by logistic regression analysis.Results:TT genotype frequency and T allele frequency of MTHFR C677T in the DVT group were higher than those in the non-DVT group: 65.00% (13/20) vs. 25.00% (20/80), 80.00% (32/40) vs. 38.75% (62/160). CC genotype frequency and C allele frequency were lower than those in the non-DVT group: 5.00% (1/20) vs. 47.50% (38/80), 20.00% (8/40) vs. 61.25% (98/160), with statistically significant differences ( P<0.05). There was no significant difference in CT genotype frequency between the two groups ( P>0.05). 4G/4G gene frequency and 4G allele frequency of PAI-1 gene in the DVT group were higher than those in the non-DVT group: 50.00% (10/20) vs. 21.25% (17/80), 67.50% (27/40) vs. 38.75% (62/160). 5G/5G gene frequency and 5G allele frequency were lower than those in the non-DVT group: 15.00% (3/20) vs. 43.75% (35/80), 32.50% (13/40) vs. 61.25% (98/160), with statistically significant differences ( P<0.05). There was no significant difference in 4G/5G gene frequency between the two groups ( P>0.05). The distribution frequency of TT genotype of MTHFR C677T and 4G/4G genotype of PAI-1 promoter in DVT group was higher than that in non-DVT group: 55.00% (11/20) vs. 22.50% (18/80), with a statistically significant difference ( P<0.05). Multivariate Logistic regression analysis showed that MTHFR C677T TT genotype ( OR = 1.499, 95% CI 1.201 to 1.871), PAI-1 promoter 4G/4G genotype ( OR = 1.471, 95% CI 1.170 to 1.850) and MTHFR The C677T loci TT genotype combined with the 4G/4G genotype of the PAI-1 promoter ( OR = 1.592, 95% CI 1.258 to 2.014) were risk factors for lower limb DVT after colorectal cancer surgery ( P<0.05). Conclusions:The TT genotype of MTHFR C677T site and the 4G/4G genotype of PAI-1 promoter are closely related to the formation of lower limb DVT after colorectal cancer surgery, and the risk of lower limb DVT is higher in patients with both genotype TT and 4G/4G.

BACKGROUND: Lung cancer is the cancer with the highest morbidity and mortality at home and abroad at present. Using computed tomography (CT) to screen lung cancer nodules is a huge workload. To test the effect of artificial intelligence in automatic identification of lung cancer by using artificial intelligence to find the lung cancer nodules automatically in the chest CT of 1 mm and 5 mm thick. METHODS: 5,000 cases of T1 stage lung cancer patients with 1 mm and 5 mm layer thickness were respectively labeled and learned by computer neural network, the algorithm of forming pulmonary nodules was carried out. 500 cases of chest CT in T1 stage lung cancer patients with 1 mm and 5 mm thickness were tested by artificial intelligence formation, and the sensitivity and specificity were compared with artificial reading. RESULTS: Using artificial intelligence to read chest CT 500 in 5 mm, the sensitivity was 95.20%, the specificity was 93.20%, and the Kappa value of two times repeated read was 0.926,1. For 1 mm chest CT 500 cases, the sensitivity is 96.40%, the specificity is 95.60%, and the Kappa reads two times is 0.938,6. Compared with 5 doctors, the same CT sets with 1 mm thickness were read. The detection rates of artificial intelligence and artificial reading were similar to those of lung cancer nodules and negative control read films, and there was no significant difference between them. In the comparison of the same CT slices with 5 mm thickness, the number of detection of lung cancer nodules by artificial intelligence is better than that of artificial reading, and the sensitivity is higher, but the number of false messages is increased and the specificity is slightly worse. CONCLUSIONS The automatic learning of early lung cancer chest CT images by artificial intelligence can achieve high sensitivity and specificity of early lung cancer identification, and assist doctors in the diagnosis of lung cancer.
Artificial Intelligence ; Humans ; Lung Neoplasms ; diagnosis ; pathology ; Medical Informatics ; methods ; Neoplasm Staging

Objective: To investigate the effect of three-dimensional conformal radiotherapy (3D-CRT) combined with PC chemotherapy (paclitaxel + carboplatin) on non-small cell lung cancer (NSCLC) patients and the serum levels of CA125, tissue inhibitor of metalloproteinase-1 (TIMP-1), serum amyloid A (SAA) and T-lymphocyte subsets. Methods: A total of 100 patients with NSCLC treated in Affiliated Hospital of Guangdong Medical University from May 2015 to December 2017 were selected as the study subjects. They were divided into control group and observation group according to random number table method, with 50 cases in each group. The observation group was treated with 3D-CRT combined with PC chemotherapy, while the control group was treated with PC chemotherapy. The two groups were treated for 4 cycles. The therapeutic effect, serum CA125, TIMP-1, SAA, T-lymphocyte subsets and adverse reactions were compared between the two groups. Results: Four cases were lost to follow-up both in the two groups. The overall response rate in the observation group (43.48%, 20/46) was higher than that in the control group (23.91%, 11/46; χ²=3.941, P=0.047). The serum levels of CA125, TIMP-1 and SAA of the two groups had no significant difference before treatment, and the levels of these indexes decreased after treatment. The serum levels of CA125, TIMP-1 and SAA in the observation group after treatment were (12.31±1.13) U/ml, (275.31±13.69) pg/ml and (47.21±7.21) mg/L, which were lower than those in the control group [(30.36±1.98) U/ml, (320.36±17.23) pg/ml, (65.92±8.36) mg/L], with significant differences (t=53.699, P<0.001; t=13.884, P<0.001; t=11.495, P<0.001). The levels of CD3⁺ , CD4⁺ , CD8⁺ and CD4⁺ /CD8⁺ of the two groups had no significant difference before treatment, and the levels of these indexes decreased after treatment. The levels of CD3⁺ , CD4⁺ , CD8⁺ and CD4⁺ /CD8⁺ in the observation group were (35.27±10.31)%, (20.27±6.72)%, (15.89±3.37)% and 0.91±0.37, which were higher than those in the control group [(30.77±9.27)%, (15.27±5.73)%, (12.02±2.69)% and 0.75±0.39], with significant differences (t=2.201, P=0.030; t=3.840, P<0.001; t=6.087, P<0.001; t=2.019, P=0.047). There were no significant differences in the adverse reactions such as nausea and vomiting [63.04% (29/46) vs. 43.48% (20/46); χ²=3.537, P=0.060], phlebitis [6.52% (3/46) vs. 4.35% (2/46); χ²=0.000, P>0.999], abnormal liver function [6.52% (3/46) vs. 2.17% (1/46); χ²=0.261, P=0.609] and myelosuppression [8.70% (4/46) vs. 6.52% (3/46); χ²=0.000, P>0.999] between the observation group and the control group. Conclusion For patients with NSCLC, 3D-CRT combined with PC chemotherapy can improve the overall response rate, decrease the levels of serum CA125, TIMP-1 and SAA, and improve the immune function of patients. The therapeutic effect is remarkable and the safety is good. The therapeutic scheme is suitable for the treatment of NSCLC.

Objective To investigate the change in body weight over time in rectal cancer patients receiving radiotherapy and the correlation between setup errors and weight loss,and to establish the image-guided radiotherapy regimens in different periods of treatment.Methods A total of 24 postoperative patients with rectal cancer admitted to our hospital in 2016 were selected.Before each fraction of radiotherapy,the body weight was recorded,and the patients underwent cone-beam computed tomography (CBCT) with different frequencies in every week.The planning CT was matched with CBCT to obtain setup errors.The paired t test was used for difference analysis;the Pearson method was used to analyze the correlation between setup errors and weight loss.Results Body weight was measured 456 times in the 24 patients,and these patients underwent CBCT scans and image registration 456 times.Two patients were excluded because of treatment discontinuance.In the first and second weeks,there was no significant change in body weight.In the third week,the mean weight loss was 1.53 kg.In the fourth week,the mean weight loss was 2.48 kg.In the fifth week,the mean weight loss was 3.24 kg.The setup errors obtained by CBCT image registration in the superior-inferior (SI),anterior-posterior (AP),and left-right (LR) directions were 0.19 cm,0.20 cm,and 0.18 cm,respectively,in the first week,0.18 cm,0.17 cm,and 0.15 cm,respectively,in the second week,0.20 cm,0.22 cm,and 0.21 cm,respectively,in the third week,0.19 cm,0.25 cm,0.24 cm,respectively,in the fourth week,and 0.34 cm,0.33 cm,and 0.31 cm,respectively,in the fifth week.The Pearson correlation analysis showed that weight loss increased the setup errors,with P values of 0.140,0.046,and 0.044 in the SI,AP,and LR directions,respectively.Conclusions For rectal cancer patients receiving radiotherapy,the body weight decreases significantly in the late period (especially in the fifth week),which influences the setup errors.Therefore,in the fourth and fifth weeks of radiotherapy for rectal cancer,the weight loss should be closely monitored,and the number of CBCT scans can be increased before the treatment fraction to ensure the accuracy and optimization of treatment.

Objective To purify pre-ribosome and ribosome of mammalian ceils using continuous sucrose density gradient ultracentrifugation.Methods Continuous sucrose density gradient was established by ultracentrifugation,and the continuous sucrose density gradient of 10％-30％ and 10％-45％ were used to extract the pre-ribosome and ribosome in mammalian cells,respectively.The mammalian cell lysis buffer was added to the established continuous sucrose density gradient.Pre-ribosome and ribosome with different sedimentation coefficients were collected and the A260 absorbance of each sample was measured.Proteins of each sample were extracted to detect the large subunit protein,RPL15 by Western Blot.Results Large subunit ribosomal protein RPL15 exists on 60S of the pre-ribosome,and also on 60S,80S and polyribosome of mature ribosome.Conclusions The continuous sucrose density gradient,which is established by the swing-out rotor,can be used to isolate the pre-ribosome and ribosome of mammalian cells rapidly.This method has the advantages of good separation effect and simple operation,which provides a good method for rapid and large amount preparation and separation of various kinds of ribosomes.

Objective To investigate the process that chromosome kinesin KIF4A promote cisplatin resistance in lung cancer cells.Methods Reverse transcription PCR (RT-PCR) and Western Blot experiments were performed to analyze the expression of KIF4A in lung cancer cells A549 and cisplatin (DDP) resistant cells A549/ DDP.Cell transfection, RNA interference (RNAi) experiments and thiazolyl blue tetrazolium bromide (MTT) assays were carried out to examine cell proliferation of A549 cells with overexpression of exogenous KIF4A and A549/DDP cells with depletion of endogenous KIF4A after cisplatin treatment.Results Expression of KIF4A in A549/DDP cells was higher than that in A549 cells.With overexpression of exogenous KIF4A, A549 cells displayed drug resistance to cisplatin.On the contrary, depletion of endogenous KIF4A in A549/DDP cells resulted in cisplatin sensitivity.Conclusions Chromosome kinesin KIF4A involves in the regulation of cisplatin resistance in lung cancer cells and KIF4A may be a potential and effective new biological target for treatment of lung cancer cisplatin resistance.

Objective:To explore the changes and clinical significance of serum amyloid A (SAA)level in patients with acute exacerbation of chronic obstructive pulmonary disease (AECOPD) .Methods:A total of 140 patients with AECOPD ,80 patients with stable stage COPD and 40 health controls during Jun .2012 and Dec .2013 were collected .The serum levels of SAA ,C‐reactive protein(CRP) ,tumor necrosis factor‐α(TNF‐α) and interleukin‐8 (IL‐8) were measured .Results:The serum levels of SAA ,CRP ,TNF‐α and IL‐8 in all patients with COPD were significantly higher than those in the controls (P<0 .05) .The serum levels of SAA ,CRP ,TNF‐α and IL‐8 in patients with AECOPD were significantly higher than those in patients with stable stage COPD(P<005) .SAA level was significantly correlated with TNF‐α level and IL‐8 level(r=0 .78 , r=0 .69 ,P<0 .01) .Serum CRP level was also significantly correlated with TNF‐αlevel and IL‐8 level(r=0 .68 ,r=0 .62 ,P<0 .01) .The area under ROC curve of SAA (0 .841) was larger than that of CRP (0 .749) ,and the difference was statistically significant(P<0 .05) .Conclusions:SAA could be applied as a new biomarker for AECOPD .Its serum level was correlated with the severity of disease .Early detection of SAA may be conducive to the evaluation of disease situation and the treatment strategy for disease .Thus ,it is worthy of clinical application .

OBJECTIVETo investigate the effect of poly-ADP-ribosylation in hexavalent chromium Cr(VI) induced cell damage.

METHODSThe study object, poly (ADP-ribose) glycohydrolase (PARG) deficient human bronchial epithelial cells (16HBE cells), was constructed previously by our research group. Normal 16HBE cells and PARG-deficient cells were treated with different doses of Cr (VI) for 24 h to compare the differences to Cr (VI) toxicity, meanwhile set up the solvent control group. On this basis, 5.0 µmol/L of Cr (VI) was selected as the exposure dose, after the exposure treatment, total proteins of both cells were extracted for two dimension fluorescence difference gel electrophoresis (2D-DIGE) separation, statistically significant differential protein spots were screened and identified by matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS/MS), and further validated by Western blot.

RESULTSAfter Cr (VI) treatment, the survival rate of PARG-deficient cells was higher than normal 16HBE cells. When the doses reached up to 5.0 µmol/L, the survival rate of 16HBE cells and PARG-deficient cells were respectively (59.67 ± 6.43)% and (82.00 ± 6.25)%, the difference between which was significant (t = -4.32, P < 0.05). 18 protein spots were selected and successfully identified after 2D-DIGE comparison of differential proteins between normal 16HBE cells and PARG-deficient cells before and after exposure. The function of those proteins was involved in the maintenance of cell shape, energy metabolism, DNA damage repair and regulation of gene expression. The differential expression of cofilin-1 was successfully validated by Western blot. The expression level of cofilin-1 in the 16HBE cells increased after Cr (VI) exposure with the relative expression quantity of 1.41 ± 0.04 in treated group and 1.00 ± 0.01 in control group, the difference of which was statistically significant (t = -18.00, P < 0.05), while the expression level in PARG-deficient cells had no statistically significant difference (t = -8.61, P > 0.05).

CONCLUSIONMost of the identified differential proteins are closely related to tumorigenesis, suggesting that poly-ADP-ribosylation reaction may resist the cytotoxicity of Cr(VI) by inhibiting Cr (VI) induced tumorigenesis, which provides important reference data to clarify the mechanisms of poly-ADP-ribosylation in Cr (VI) induced cell damage.

Bronchi ; Cell Transformation, Neoplastic ; genetics ; Chromium ; Cofilin 1 ; DNA Repair ; Epithelial Cells ; Glycoside Hydrolases ; deficiency ; physiology ; Humans ; Tandem Mass Spectrometry

OBJECTIVETo reveal the role of poly-ADP-ribosylation and DNA methylation in carcinogenic process induced induced by Cr (VI), and to discuss the relations between them.

METHODSThe pre-established Poly (ADP-ribose) glycohydrolase (PARG) deficient cells and 16HBE cells were treated with different concentrations of Cr (VI), and the changes of total genomic DNA methylation level in different groups were detected by methylation immunofluorescent detection, as well as the changes of the activity of methyltransferases. Moreover, RT-PCR and western blotting method were applied to analyze the changes of expression of DNMT1, DNMT3a, DNMT3b and MBD2, upon the protein level.

RESULTSAfter treated by Cr(VI) for 24 h, the healthy 16HBE cells showed a significant lower level of genomic DNA methylation; however, there was no significant changes (P > 0.05) found in PARG deficient cells by immunofluorescence assay. When the dose of Cr (VI) reached 5.0 µmol/L, the activity of methyltransferases in 16HBE cells and PARG deficient cells (49.33 ± 2.65, 80.05 ± 2.05) decreased by 20% and 50% comparing with contrast group (99.27 ± 1.10, 99.30 ± 0.60) . After treated by Cr (VI) for 24 h, the expression of mRNA and protein level among DNMT1, DNMT3a, DNMT3b and MBD2 decreased significantly in healthy 16HBE cells; and the expression of DNMT1 and DNMT3a decreased in PARG deficiency cells. The relevant expression levels of mRNA of DNMT1 were separately (0.99 ± 0.09), (0.79 ± 0.10), (0.59 ± 0.13) and (0.39 ± 0.02) (F = 247.17, P < 0.01), the expression levels of protein were separately (1.00 ± 0.03), (0.69 ± 0.15), (0.65 ± 0.10) and (0.55 ± 0.13) (F = 214.12, P < 0.01), the expression levels of DNMT3a mRNA were separately (1.00 ± 0.04) , (0.93 ± 0.11) , (0.79 ± 0.07) , (0.59 ± 0.05) (F = 498.16, P < 0.01) , and the expression levels of protein were separately (1.00 ± 0.14) , (0.97 ± 0.11) , (0.79 ± 0.17) , (0.57 ± 0.15) (F = 390.11, P < 0.01) when the dose of Cr (VI) at 0, 0.3, 1.2 and 5.0 µmol/L. However, there were no significant changes of expression found in DNMT3b and MBD2.

CONCLUSIONPoly-ADP-ribosylation could regulate the activity of DNMT3b and MBD2, protect cells against the DNA methylation alteration induced by Cr(VI) and maintain the global genomic DNA methylation level.

Cell Line ; Chromium ; toxicity ; DNA (Cytosine-5-)-Methyltransferase 1 ; DNA (Cytosine-5-)-Methyltransferases ; metabolism ; DNA Methylation ; drug effects ; DNA-Binding Proteins ; metabolism ; Epithelial Cells ; metabolism ; Genome ; Humans ; Poly Adenosine Diphosphate Ribose ; metabolism ; RNA, Messenger ; genetics