To establish a method of directional differentiation and efficient production of neurons from embryonic stem cells (ES cells) in vitro, based on the 4-/4+ protocol described by Bain, a new method was established to induce ES cells differentiating into neurons by means of three-step differentiation using all-trans retinoic acid (ATRA) combined with astrocyte-conditioned medium (ACM) in Vitro. The totipotency of ES cells was identified by observation of cells' morphology and formations of teratoma in immunocompromised mice. The cells' differentiation was evaluated continuously by the detection of the specific cellular markers of neural stem cells, neurons and astrocytes, including nestin, NSE and GFAP using immunohistochemistry assay. The NSE positive cells' ratio of the differentiated cells was determined by flow cytometry. It was found that the transparent circular clusters surrounding embryoid bodies induced with combining induction protocol formed just after 24 h and gradually enlarged later. This phenomenon could not be observed in EBs induced only by ATRA. The NSE positive cells' ratio in the cells induced with ATRA and ACM was higher than that of the cells induced by ATRA at different time points of differentiation, and finally reached up to 73.5% among the total differentiated population. It was concluded that ES cells could be induced into neurons with high purity and yield by means of inducing method combining with ATRA and ACM.
Astrocytes/*cytology ; Cell Differentiation ; Cells, Cultured ; Embryo, Mammalian ; Neurons/*cytology ; Stem Cells/*cytology ; Tretinoin/pharmacology

Objective To investigate the differences in the gene expression profiles between SW480 and SW620 cell lines.Methods A dataset of GDS756 containing the gene expression profiles of SW480 and SW620 was downloaded from the GEO database in NCBI.The differential expression genes between SW480 and SW620 were analyzed with gene set enrichment analysis (GSEA) and leading edge subset analysis.The genes in leading edge subset were re-annotated by FunRich software.The core genes of leading edge subset closely relating to SW480 or SW620 were analyzed with the STRING on-line analytical system.The functional core genes closely relating to SW480 or SW620 were obtained by the combined analysis of the core genes and high frequency genes from leading edge subset.Results GSEA identified 12 significantly enriched gene sets,491 leading edge genes and 7 highly overlapping genes from SW480 and 80 significantly enriched gene sets,870 leading edge genes and 6 highly overlapping genes from SW620.The STRING system identified 5 core genes from SW480 and 8 from SW620.The combined analysis of GSEA and bionetwork obtained 2 functional core genes,TOP2A and CDK1,from SW620.Conclusion The SW480 and SW620 cells with identical genetic background have different functional gene expression profiles,and the functional core genes TOP2A and CDK1 in SW620 cells may be related to the signal pathways of colon cancer metastasis.

Objective To investigate the expression of specific marker molecules in hair-inducing activity of long-term cultured human dermal papilla cells (HDPCs) in vitro.Methods After dissected and cultured the HDPCs in vitro,the cells of passages 1 to 8 were used for experiments.The growth appearances of HDPCs in different passages were observed under inverted microscope.To detect the expression of specific marker molecules of long-term cultured HDPCs,the alkaline phosphatase (ALP) activity of the HDPCs was examined,and the specific genes ALP and insulin-like growth factor-1 (IGF-1) expression levels of HDPCs were determined by real-time quantitative PCR.Results After long-term cultured in vitro,the ALP and IGF-1 expression levels of HDPCs gradually decreased in different passages,as well as the display of the aggregated and cartouche growth.The ALP and IGF-1 expression levels of HDPCs in passage 1 was the highest,they were almost about 6.8-fold and 3.5-fold higher than the HDPCs in passage 8.The ALP staining of the HDPCs in passage 1 and passage 2 were evident,but the cells' ALP staining gradually became much weaker than the cells in the previous passages after the long-term cultured in vitro.Conclusions The expression levels of specific marker molecules ALP and IGF-1 of the HDPCs decrease gradually after long-term cultured in vitro,and the higher passage HDPCs lost the special aggregated and cartouche growth appearance,and hence lead to the loss of hair-inducing activity of HDPCs.

Objective To study the long-term effect of administration(6 months) with transient receptor potential vanilloid 1(TRPV1) agonist capsaicin on contractile reactivity of thoracic aorta in C57BL/6J mice.Methods Tow-month-old male C57BL/6J mice were received normal diet group(n=12) or capsaicin group(normal diet plus capsaicin,n=12).Tail-cuff systolic blood pressure(SBP) was examined at the baseline and at the end of the intervention.After 6-month treatment period,carotid artery blood pressure and heart rate were determined by catheterization,and the aortic contractile response was examined using isometric myograph(Danish Myotech Technology,Denmark).Plasma levels of renin,angiotensin Ⅱ(Ang Ⅱ) and aldosterone were determined.Vascular smooth muscle cells(VSMC) were obtained from thoracic aorta of mice and cultured.Angiotensin Ⅱ type 1 receptor(AT1R) protein expression was detected by western blot.Calcium imaging was detected in cultured VSMC using the fluorescent dye technique.Results Systolic blood pressure,invasive carotid artery blood pressure and heart rate have no difference between two groups.No differences was found in PE-induced contraction response in thoracic aorta;while Ang Ⅱ induced contractility of aortic ring was lower in mice with capsaicin than control group [capsaicin:(37.5?1.6)% vs(59.8?1.4)%,P

To establish a method of directional differentiation and efficient production of neurons from embryonic stem cells (ES cells) in vitro, based on the 4-/4+ protocol described by Bain, a new method was established to induce ES cells differentiating into neurons by means of three-step differentiation using all-trans retinoic acid (ATRA) combined with astrocyte-conditioned medium (ACM) in Vitro. The totipotency of ES cells was identified by observation of cells' morphology and formations of teratoma in immunocompromised mice. The cells' differentiation was evaluated continuously by the detection of the specific cellular markers of neural stem ceils, neurons and astrocytes,including nestin, NSE and GFAP using immunohistochemistry assay. The NSE positive cells' ratio of the differentiated cells was determined by flow cytometry. It was found that the transparent circular clusters surrounding embryoid bodies induced with combining induction protocol formed just after 24 h and gradually enlarged later. This phenomenon could not be observed in EBs induced only by ATRA. The NSE positive cells' ratio irthe cells induced with ATRA and ACM was higher than that of the cells induced by ATRA at different time points of differentiation, and finally reached up to 73.5 % among the total differentiated population. It was concluded that ES cells could be induced into neurons with high purity and yield by means of inducing method combining with ATRA and ACM.

OBJECTIVEThe effect of allitridin on the transcription levels of immediate-early (ie), early(e) and late (1) genes of human cytomegalovirus (HCMV) was investigated in order to explore the mechanism of allitridin against HCMV.

METHODEstablished the models of HCMV AD169 strain infected cells and AD169 strain infected cells treated with allitridin (9.6 mg x L(-1)), and they were compared with the appropriate dose(2.3 mg x L(-1)) of ganciclovir (GCV). All groups of cells were infected at 2.5 multiplicity of infection (MOI), using SYBR Green real-time PCR method to detect the dynamic change of ul122, ul123, ul54 and ul83 mRNA expression at 0.5, 2, 4, 6, 12, 24 h post-infection.

RESULTThe mRNA levels of ul122 and ul123 in AD169 infected cells treated with allitridin at all time points were markedly lower than those of AD169 infected controls (P<0.05), but there were no significant difference of ul122 gene in AD169 infected cells treated with GCV and AD169 infected cells at 0.5-6 h post-infection. The inhibitory rates of allitridin to AD169 ul122 and ul123 mRNA reached 75.2% and 70.4% at 24 h post-infection, respectively. The expression of ul54 mRNA in two drug-treatment groups at all time points were lower than those of AD169 infected cells group (P<0.05). The inhibitory rates of alltridin and GCV to AD169 ul54 mRNA were 45.4% and 27.2% at 24 h post-infection,respectively. The expression of HCMV ul83 mRNA in all groups rapidly increased after 6 h of infection,which is most obvious in AD169 infected cells group. The inhibitory rates of alltridin and GCV to AD169 ul83 mRNA were 45.9% and 26.2% at 24 h post-infection, respectively.

CONCLUSIONAllitridin could effectively suppress the transcription of ie genes (ul122 and ul123) of HCMV AD169 strain, led the expression of mRNA significantly lowerd. It was able to supress the transcription of egene (ul54) and l gene (ul83) too, indicating that HCMV ie genes may be the key target of allitridin against HCMV.

Allyl Compounds ; pharmacology ; Antiviral Agents ; pharmacology ; Cell Line ; Cytomegalovirus ; drug effects ; genetics ; Genes, Immediate-Early ; genetics ; Genes, Viral ; genetics ; Humans ; Sulfides ; pharmacology ; Transcription, Genetic ; drug effects

Objective:To observe the clinical effects of autologous fat transplantation combined with liposuction in buttock shaping.Methods:January 2020 to January 2021, fifty-eight female patients (aged 21-36 years) with hip depression, hip prolapse, obvious inferior plica and fat hypertrophy of buttocks were treated in the clinic of medical cosmetology of Hangzhou Tongrong Lige Medical Technology Co., Ltd., the average age is 29. There were 20 cases of waist-abdomen liposuction + hip shaping, 18 cases of thigh liposuction + hip shaping, 16 cases of thigh liposuction + hip shaping, and 4 cases of simple hip shaping. All patients were followed up for 3-12 months to observe the shape of buttocks and complications (hematoma, infection, fat liquefaction).Results:A total of 58 patients were followed up for 3-12 months. Among the 58 patiens, 45 patiens had significant effect, 11 patients had good effect, and 2 patients had poor effect. Of the 11 cases with good results, 6 cases had poor hip shape and 5 cases had slight symmetry of the lateral buttocks. The reason was that the patients′ buttocks were flat and needed 2-3 times of filling. The buttocks were filled again one year after operation, and better results were achieved. No complications such as infection, hemmatoma, fat liquefaction and uneven bump occurred.Conclusions:The patients are satisfied with the shape of the buttocks after autologous fat transplantation combined with liposuction in buttock shaping. The incidence of surgical complications is low.