Establishment of HPLC fingerprint and content determination of multiple index components in Xiao’er resuqing oral liquid / 中国药房

OBJECTIVE To establish an HPLC fingerprint of Xiao’er resuqing oral liquid， and to determine the contents of twelve index components. METHODS HPLC method was adopted. The determination was performed on Venusil MP C18 column with mobile phase consisting of acetonitrile-0.1% phosphate aqueous solution （gradient elution） at a flow rate of 1.0 mL/min. The detection wavelength was set at 210 nm， the column temperature was 30 ℃， the injection volume was 10 μL. HPLC fingerprint of Xiao’er resuqing oral liquid was established by using the Similarity Evaluation System of Chromatographic Fingerprint of TCM （2012 edition） to evaluate the similarity. The contents of 12 components were determined， including （R， S）-goitrin， 3，5-O-dicaffeoyl quinic acid， puerarin， forsythin， forsythoside A， chlorogenic acid， baicalin， saikosaponins d， wogonoside， baicalein， emodin and chrysophanol. RESULTS The similarity of HPLC fingerprints of 13 batches of Xiao’er resuqing oral liquid was greater than 0.97， and 14 common peaks were confirmed. The contents of the above 12 index components in 13 batches of Xiao’er resuqing oral liquid were as follows： 0.078-0.172， 1.564-2.736， 1.338-2.578， 0.426-0.872， 1.477-2.628， 1.396-2.447， 4.052-9.146， 0.367- 0.692， 1.974-4.674， 1.274-2.969， 0.085-0.167 and 0.155-0.307 mg/mL. CONCLUSIONS The established HPLC fingerprint and content determination methods have high accuracy and high specificity， which can be used for the quality evaluation of Xiao’er resuqing oral liquid.

Compliance with retainer wear using audiovisual integration and reminder: a randomized clinical trial.

Active audiovisual representation of instructions ensures vibrant knowledge acquisition and improves acquaintance needed for self-care with retainer wear. The aim of this trial is to assess the impact of audiovisual instructions with additional weekly electronic reminder messages on improving adherence to instructed wear time of Hawley retainer, periodontal outcomes, and participants' experiences. Fifty-two participants (mean age 26.1 y) planned for removable retention, were randomly assigned to two parallel groups to receive either (1) audiovisual instructions with an additional weekly reminder, or (2) verbal instructions alone. Each participant received a Hawley retainer equipped with a TheraMon microsensor and was instructed to wear it for 22 h daily. Participants were monitored for adherence to the wear time after 3 (T1) and 6 months (T2), and had their periodontal health and experiences assessed at T2. Overall, the mean objectively measured daily wear time at T1 was 14.9 (± 4.9 h), and 14.3 (± 5.4 h) at T2. After 3 months, no significant differences were found between the groups (p = 0.065), however, a significant difference favoring better compliance with wear instructions was observed in the audiovisual group after 6 months (p = 0.033). A non-significant difference was observed between both groups regarding the gingival (p = 0.165) and plaque index scores (p = 0.173). Participants' experiences were similar in both groups, except for satisfaction with the way of delivering instructions, being favorably reported in the audiovisual group. Audiovisual instructions with weekly reminders seem to have a significant effect on patient compliance in the longer term.Trial registration: TCTR20230220002.

Effect of chlorhexidine mouthwash on gingival health around orthodontic miniscrew implants: A pilot placebo-controlled randomized trial.

OBJECTIVE: This randomized controlled trial (RCT) aims to investigate the short-term effects of chlorhexidine mouthwash (MW) on gingival health surrounding orthodontic miniscrew implants (OMIs) and their overall survivability. MATERIALS AND METHODS: Thirty-two participants (mean age, 22.8 years) undergoing fixed orthodontic appliance treatment after maxillary premolar extraction were randomly allocated in a parallel fashion to either receive (1) MW with an active component of chlorhexidine or (2) a placebo. Each participant received two maxillary buccal OMIs for anchorage reinforcement purposes. Participants were assessed for their gingival oral health status around all inserted OMIs and had their OMI survivability recorded at three time points; T1 = 1 month, T2 = 3 months, and T3 = 6 months after OMI placement. A Kaplan-Meier plot was used to estimate the survival function of OMIs. RESULTS: All randomized participants completed the follow-up period. In terms of gingival oral health, there were no statistically significant differences at any time point between the chlorhexidine MW group and the placebo-controlled group (P > .05). One OMI was lost in the chlorhexidine MW group and another two OMIs in the control group. There was no significant difference between both groups in terms of survivability (P = .585). CONCLUSION: The use of chlorhexidine MW does not seem to have a significant clinical impact on gingival health around OMIs or their survivability in this pilot study.

Progress in molecular signal pathway of dental fluorosis / 中华地方病学杂志

Dental fluorosis is a manifestation of chronic oral fluorosis caused by excessive fluoride intake in childhood. At present, the molecular mechanism of dental fluorosis is still unclear. Ameloblasts are the most sensitive cells to fluoride in tooth tissue. Fluoride affects the proliferation and secretion of ameloblasts through the role of key molecules in the molecular signal pathways, leading to the formation of dental fluorosis. This paper reviews the relationship between the molecular signal pathways [mitogen-activated protein kinase (MAPK), transforming growth factor-β (TGF-β)/Smad, Wnt, Foxo1/Runx2], stress pathways (endoplasmic reticulum stress and oxidative stress) and the occurrence of dental fluorosis in recent years, in order to deeply understand the pathogenesis of dental fluorosis at the molecular level, and provide new ideas for the prevention and treatment of dental fluorosis.

CRISPR/Cas9-mediated Knockout of KIFC1 Inhibits Proliferation and Induces Apoptosis of Cervical Cancer Cells / 肿瘤防治研究

Objective To investigate the functions of the KIFC1 gene in tumor cells and its effect on the proliferation of cervical cancer cells. Methods We designed sgRNAs targeting the KIFC1 gene and constructed a recombinant plasmid based on the pSpCas9 (BB)-2A-GFP vector, which was co-transfected into HeLa cells. We screened monoclonal knockout cell lines through flow cytometry sorting, limited dilution inoculation of cells, and sequencing. RT-qPCR, Western blot, and immunofluorescence were used to detect the transcription and protein expression levels of KIFC1 in knockout cells. Cell phenotypes such as nucleus and microtubule cytoskeleton were observed using phase-contrast microscopy and fluorescence confocal microscopy. Cell proliferation, cell cycle, and apoptosis were analyzed by growth curve plotting, EdU labeling, and acridine orange staining. Results The deletion of the KIFC1 gene resulted in the abnormal phenotypes of HeLa cells, with increased numbers of multinuclei, micronucleus, and disordered microtubules. The cell cycle was disrupted, accompanied with a significant increase in the ratio of late apoptotic cells and a decrease in cell proliferation (all P < 0.05). Conclusion KIFC1 gene deletion affects the assembly of microtubules and cell division in HeLa cells, leading to abnormal nuclear morphology, chromatin elimination, cell cycle arrest, and increased cell apoptosis.

Identification of a NONO gene variant in a child with congenital heart disease and global developmental delay / 中华医学遗传学杂志

OBJECTIVE@#To explore the genetic basis for a child with congenital heart disease (CHD) and global developmental delay (GDD).@*METHODS@#A child who was hospitalized at the Department of Cardiac Surgery of Fujian Children's Hospital on April 27, 2022 was selected as the study subject. Clinical data of the child was collected. Umbilical cord blood sample of the child and peripheral blood samples of his parents were collected and subjected to whole exome sequencing (WES). Candidate variant was verified by Sanger sequencing and bioinformatic analysis.@*RESULTS@#The child, a 3-year-and-3-month-old boy, had manifested cardiac abnormalities and developmental delay. WES revealed that he had harbored a nonsense variant of c.457C>T (p.Arg153*) in the NONO gene. Sanger sequencing showed that neither of his parents has carried the same variant. The variant has been recorded by the OMIM, ClinVar and HGMD databases, but not in the normal population databases of 1000 Genomes, dbSNP and gnomAD. Based on the guidelines from the American College of Medical Genetics and Genomics (ACMG), it was rated as a pathogenic variant.@*CONCLUSION@#The c.457C>T (p.Arg153*) variant of the NONO gene probably underlay the CHD and GDD in this child. Above finding has expanded the phenotypic spectrum of the NONO gene and provided a reference for the clinical diagnosis and genetic counseling for this family.

Lineage-mosaic and mutation-patched spike proteins for broad-spectrum COVID-19 vaccine

The widespread SARS-CoV-2 in humans results in the continuous emergence of new variants. Recently emerged Omicron variant with multiple spike mutations sharply increases the risk of breakthrough infection or reinfection, highlighting the urgent need for new vaccines with broad-spectrum antigenic coverage. Using inter-lineage chimera and mutation patch strategies, we engineered a recombinant monomeric spike variant (STFK1628x), which showed high immunogenicity and mutually complementary antigenicity to its prototypic form (STFK). In hamsters, a bivalent vaccine comprised of STFK and STFK1628x elicited high titers of broad-spectrum antibodies to neutralize all 14 circulating SARS-CoV-2 variants, including Omicron; and fully protected vaccinees from intranasal SARS-CoV-2 challenges of either the ancestral strain or immune-evasive Beta variant. Strikingly, the vaccination of hamsters with the bivalent vaccine completely blocked the within-cage virus transmission to unvaccinated sentinels, for either the ancestral SARS-CoV-2 or Beta variant. Thus, our study provides new insights and antigen candidates for developing next-generation COVID-19 vaccines.

Advances and applications of organoids: a review / 生物工程学报

Novel model systems have provided powerful tools for the research of human biology. Despite of being widely used, the conventional research models could not precisely describe the human physiological phenomenon. Organoids are three-dimensional multicellular aggregates derived from stem cells or organ progenitors that could differentiate and self-organize to recapitulate some specific functionalities and architectures of their in vivo counterpart organs. Organoids can be used to simulate organogenesis because of their human origin. In addition, the genomic stability of organoids could be well maintained during long-term amplification in vitro. Moreover, organoids can be cryopreserved as a live biobank for high-throughput screening. Combinatorial use of organoids with other emerging technologies (e.g. gene editing, organ-on-a-chip and single-cell RNA sequencing) could overcome the bottlenecks of conventional models and provide valuable information for disease modelling, pharmaceutical research, precision medicine and regenerative medicine at the organ level. This review summarizes the classifications, characteristics, current applications, combined use with other technologies and future prospects of organoids.

Correlation between BMI and metagenomics concentration of gut microbiota of college students in Inner Mongolia / 中国学校卫生

Objective@#To explore the correlation between BMI and gut microbiota of college students in Inner Mongolia,and to provide a reference basis for revealing the relationship between intestinal flora and obesity.@*Methods@#Totally 88 college students from Inner Mongolia Medical University were enrolled, Height and weight were measured,and the feces samples were collected. The bacterial metagenome was extracted from dry feces samples for the concentration detection in per gram of dry feces,expressed as μg/μL. Correlation between BMI and metagenomics concentration of gut microbiota was statistically analyzed. Meanwhile,the metagenomics concentration of gut microbiota in different BMI groups was compared with each other.@*Results@#There was a negative correlation between BMI and the metagenomics concentration of gut microbiota(r=-0.27,P<0.05). Significant difference in the concentration of gut microflora was observed between the normal group and the obesity group,the normal group and the overweight/obesity group(F=3.62,P<0.05). Among the female volunteers,there were significant differences between normal group and overweight group,between normal group and obesity group(F=1.87,P<0.05). No significant differences in metagenomics concentration of gut microbiota were found in different BMI groups(F=0.60, P>0.05).@*Conclusion@#There is a correlation between BMI and gut microbiota of college students in Inner Mongolia，the concentration of gut microflora metagenome in overweight and obese people decreased significantly.

Analysis of influencing factors of the effect of the morphology of unruptured intracranial aneurysms on aneurysm wall enhancement of vascular wall with high-resolution magnetic resonance imaging / 中国脑血管病杂志

Objective To investigate the influencing factor of the morphology of unruptured intracranial aneurysms for aneurysm wall enhancement under the high-resolution magnetic resonance imaging. Methods From January 2015 to December 2016,the clinical and imaging data of 68 consecutive patients with unruptured intracranial aneurysm (86 aneurysms) in Changhai Hospital,the Second Military Medical University were enrolled retrospectively. Vascular wall imaging technology was used to conduct aneurysm scan,and the aneurysm wall enhancement was identified by the imaging features before and after contrast enhancement. They were divided into either an enhancement group ( n=32,34 aneurysms) or a non-enhancement group (n=45,52 aneurysms) according to whether having the abnormal enhancement of aneurysm wall or not ( because some patients also have enhanced aneurysms and non-enhanced aneurysms, the number of cases of the enhanced or not was calculated seperately in both groups ) . Morphological parameters were calculated by 3D image data,including aneurysm size,ratio of height to width,volume ratio, dome-to-neck ratio, transverse length ratio, bottleneck factor, and inflow angle. Univariate and multivariate logistic analyses were used to determine the morphological influence factors of aneurysm wall enhancement. Results (1) A total of 34 (39. 5%) aneurysms had aneurysm wall enhancement and 52 (60. 5％) aneurysms did not have aneurysm wall enhancement. There were no significant differences in sex, age, hypertension,diabetes, smoking, family history of subarachnoid hemorrhage, and aneurysm site in both groups (all P>0. 05). (2) The aneurysm size,ratio of height to width,volume ratio,dome-to-neck ratio, and bottleneck factor in the enhancement group were larger than those of the non-enhancement group. There were significant differences between the 2 groups (9. 19 [6. 54,11. 04] mm vs. 5. 31 [4. 17,7. 37] mm, (1. 18 [1. 01,1. 69] vs. 0. 91 [0. 72,1. 25],(3. 62 [2. 30,4. 63] vs. 2. 18 [1. 37,2. 76],1. 52 [1. 25, 1. 99] vs. 1. 19 [1. 03,1. 51],and 1. 21 [1. 11,1. 69] vs. 1. 05 [0. 94,1. 31],all P<0. 01). The proportion of irregular morphologic aneurysms in the enhancement group was higher than that in the non-enhancement group. There was significant difference between the 2 groups (55. 9％ [19/34] vs. 17. 3％ [9/52],P<0. 01 ) . There were no significant differences in transverse length ratio and inflow angle between the 2 groups (all P>0. 05). (3) Because the ratio of height to width,volume ratio,dome-to-neck ratio,and bottleneck factor were related to the aneurysm size,the aneurysm size,inflow angle,and irregular shape were included in the multivariate logistic regression analysis. The results showed that aneurysm size ( OR,3. 727,95％CI 1. 933-6. 971,P<0. 01) and irregular shape (OR,3. 990,95％CI 1. 219-13. 065,P=0. 022) were the independent risk factors for aneurysm wall enhancement. Conclusions The size and irregular shape of unruptured intracranial aneurysms are the independent risk factors for aneurysm wall enhancement. High-resolution magnetic resonance wall imaging may become an effective and noninvasive imaging method for evaluating the ruptured risk of intracranial aneurysms.

Relationship between polymorphism of surfactant protein gene C and neonatal respiratory distress syndrome / 中华实用儿科临床杂志

Objective Neonatal respiratory distress syndrome (NRDS) is one of the severe respiratory complications in the early stage after birth.NRDS can develop ingravescence respiratory difficulty due to insufficient pulmonary surfactant and its mortality is very high.This research was to study the common mutation sites of surfactant pulmonary-associated protein C (SFTPC) gene and its relationship with NRDS.Methods All 46 neonates with NRDS at the Department of Neonatology,Children's Hospital of Fudan University from January 2012 to March 2015 were assigned as case group and other 44 neonates without NRDS as control group.All cases were Han race.SP-C gene was tested with Sanger sequencing based genetic testing.All data were analyzed with SPSS 20.0 software.Results There was no significant difference in sex,gestational age,birth weight or mode of delivery between case group and control group(all P ＞ 0.05).The mutations reported in European and African neonates with respiratory distress syndrome were not found in this study.There were 3 nonsense mutations in the study group,c.-1614C ＞ A,c.-1504G ＞ A and c.-368A ＞ G,and their Regulome DB Scores were 4,5 and 5 scores.There was a significant difference in the mutation at the spot rs8192308 between the two group and their Regulome DB Scores was 5 scores.Conclusion The mutation in spot rs8192308 may have relationship with the risk of NRDS in Han race of Shanghai city.

Value of reduced field of view DWI in differentiating solid pancreatic focal lesions / 中华胰腺病杂志

Objective To study the value of reduced field-of-view (rFOV DWI) in differentiating patients with solid pancreatic focal lesions.Methods 139 patients with solid pancreatic mass were enrolled,including 105 patients with pancreatic ductal acinar carcinoma (PDAC),16 patients with neuroendocrine neoplasms,7 patients with mass forming chronic pancreatitis (MFCP) and 11 patients with solid papillary tumor (SPT).38 healthy adult volunteers served as controls,and underwent single stimulated echo planar imaging (ss-EPI) DWI and rFOV DWI(b value =0 and 600 s/mm2) MRI examination.Quartation method was used to evaluate the image quality of ss-EPI) DWI and rFOV DWI in the three terms of the visibility of anatomical structure,contrast of pancreatic lesions,motion and the susceptibility artifacts during MRI.Work station self-carried software was used to measure the ADC value of the region of interest (ROI).The image quality and ADC values of different pancreatic diseases and normal pancreas were compared.ROC curve for ADC value was drawn to evaluate the difference among PDAC,other benign pancreatic masses and normal pancreas.Results At b value of 0 and 600 s/mm2,rFOV DWI was superior to ss-EPI DWI in terms of showing pancreatic anatomic structure,the contrast of the lesion and the score evaluation for susceptibility artifacts(b =0 s/mm22.99 ±0.51 vs 2.79 ±0.64,2.37±0.48 vs 1.81 ±0.63,3.17 ±0.56 vs 2.91 ±0.60;b =600 s/mm23.63 ±0.50 vs 3.32 ±0.56,3.45 ±0.50 vs 3.01 ±0.49,3.74 ±0.44 vs 3.12 ±0.37),and the differences were statistically significant (P＜0.001).ADC values of PDAC,NET,MFCP,SPT and normal pancreas were (1.38 ± 0.17) × 10-3,(1.22 ± 0.35) × 10-3,(1.29 ± 0.13) × 10-3,(1.04 ± 0.38) ×10-3and(1.86±0.15) ×10-3mm2/sforrFOV DWI,and (1.73 ± 0.24) ×10-3,(1.63±0.39) ×10-3,(1.58±0.19) × 10-3,(1.25±0.26) × 10-3 and(2.04±0.20) × 10-3mm2/s for ss-EPI DWI.The difference on ADC values among different groups and within one group were all statistically significant (P ＜0.001).There were no statistical significant differences on ADC values between MFCP and PDAC,between MFCP and SPT as well as on ss-EPI DWI ADC values between PDAC and NET,but statistical differences were found between other two groups (P ＜ 0.05).The area under the ROC curve of rFOV and ssEPI DWI was 0.983 (95％ CI 0.944-0.998) and 0.889 (95％ CI 0.822-0.936),respectively,and the difference was statistically significant (P =0.0004),but rFOV DWI and ss-EPI DWI ADC values for PDAC and all benign solid diseases were 0.799 (95％ CI 0.719-0.864) and 0.755 (95％ CI 0.672-0.827),and the difference was not statistically significant.Conclusions rFOV DWI could significantly enhance the quality of DWI images,and its diagnostic efficacy was much better than ss-EPI DWI.

Preparation of monoclonal antibodies against ricin toxin and development of up-converting phosphor technology-based lateral flow assay for its quantitative detection / 军事医学

Objective To develop an up-converting phosphor technology based lateral flow assay ( UPT-LF) to detect ricin toxin ( RT) quickly, accurately and quantitatively.Methods Ricin-monoclonal antibodies were prepared and their affinity was evaluated before four types of monoclonal antibodies with the highest titer were applied to couple with the up-converting phosphor nano-particles ( UCP-NPs) as the bio-conjugate and disperse on the analysis membrane as the test line, respectively.Following systematic optimization to establish the RT-UPT-LF strip, the sensitivity, precision, quantita-tive ability and specificity of RT-UPT-LF were evaluated.Results The detection could be accomplished within 15 min and the detection limit of the RT-UPT-LF assay could reach 0.5 ng/ml within the quantitative detection range of 0.5-1000 ng/ml.Other non-specific toxins at a concentration of 1000 ng/ml did not cause any non-specific reactions.Conclusion The developed RT-UPT-LF strip provides a new means for on-site quantitative detection of ricin toxin.

Application of diffusion tensor imaging in the neonatal brain development and injury / 中华实用儿科临床杂志

Diffusion tensor imaging (DTI)can reflect the brain structure and development more quantitatively and intuitively than conventional magnetic resonance sequence by obtaining the diffusion properties of water molecules. In neonatal brain developmental research,DTI could be used to study the developmental regularities of white matter tracts and cerebral structure deformity.It can also help to explore the relationship between white matter microstructure and neurodevelopmental outcome.In the study of brain injury,including premature white matter injury,hypoxic ischemic encephalopathy,neonatal stroke,and so on,DTI can diagnose the brain microstructure injury precisely,evaluate the ef-fectiveness of interventions and predict the long -term neurodevelopmental outcome.DTI may have good prospect in re-search and clinical application on neonatal brain development and injury.

Clinical application of tranexamic acid in perioperative period of total knee arthroplasty / 中国组织工程研究

amount of bleeding to different degrees. It is believed that with the wide use of tranexamic acid during and after total knee arthroplasty, there wil be more optimal mode that can better control blood loss after total knee arthroplasty.

Interleukin-1β induced transdifferentiation of renal tubular epithelial cells and its effect on the cytoskeleton / 中国免疫学杂志

Objective:To investigate the effect of interleukin-1β (IL-1β) on epithelial-mesenchymal transition (EMT) and cytoskeleton rearrangement of renal tubular epithelial cells.Methods:Immortalized renal tubular epithelial cell line NRK52E was cultured in vitro with IL-1β (30 μg/L) for 3 days and 6 days,then the cell morphology was observed;The mRNA expressions of α-smooth muscle actin (α-SMA),cytoskeleton components β-actin and α-tubulin were semi-quantitative examined by RT-PCR.The protein expression of α-SMA and arrangements of β-actin and α-tubulin were assessed by immunofluorescent staining.Results:After induced by IL-1β for 3 days and 6 days in vitro,the mRNA and protein expression of α-SMA increased significantly compared with corresponding control cells (P<0.001),it prompted that NRK52E cells underwent EMT;At the same time,the cell morphology also changed,from a typical multilateral paving stone to fibroblast-like appearance,with multiple processes; Cytoskeletal protein β-actin mRNA expression was also slightly increased (P<0.05).The distributions and arrangements of β-actin protein were also changed,from cell membrane transferred to peri-nucleus and cytoplasm,moreover it formed fiber bundle-like structures.However,another cytoskeleton protein α-tubulin in IL-1β induced cells,neither it's mRNA expression nor it's distribution had significant differences compared with the control group.Conclusion:IL-1β can induce NRK52E cells undergoing EMT in vitro,cell morphology changes into fibroblast-like appearance with multiple processes,and also the cytoskeleton protein β-actin expression increases and rearrangement occurrs.However,there was no changes onα-tubulin.

Effect of glial cell-derived neurotrophic factor-modified adipose-derived stem cells on survival of co-cultured dopaminergic neurons / 中国组织工程研究

BACKGROUND: Reports regarding adipose-derived stern cells (ADSCs) differentiation into dopaminergic (DN) neurons are few in addition, there is not experimental evidence of the effect of ADSCs on maintaining the survival of DN neurons.OBJECTIVE: To investigate the effect of glial cell-derived neurokophic factor (GDNF)-modified adipose-derived stem cells on survival of DN neurons under co-cultured condition.DESIGN, TIME AND SETrlNG: The in vitro cytology experiment was conducted at the Institute of Otolaryngology-Head and Neck Surgery and Key Laboratory of Zoonoses of Ministry of Education between March and December 2007.MATERIALS: Wistar rats with 3-weeks-old, or 14 days of pregnancy were provided by Norman Bethune College of Medicine, Jilin University.METHODS: The GDNF recombinant adenovirus was constructed by using pAdTrackCMV and pAdEasy-1 system. DN neurons were obtained from the rostral mesencaphalic tegmentum of Wistar rat embryos by using trypsin and collagenase method. ADSCs isolated from rat inguinal fat pads were digested with collagenase Ⅱ, cultured and passaged in vitro. When the cells reached 60% cenfluency at the 3rd passage, cells were transfectad with 1×109vp/mL of Ad-GDNF for 1 hour and then transferred into growth medium for another 24 hours, and GDNF level in cell supematant was detected by ELISA assay. Meanwhile, the co-cultured of ADSCs and DN neurons were carried out for following 7 days. With GFP-modified ADSCs was served as a control group.MAIN OUTCOME MEASURES: The effect of co-cultured condition on the survival of DN neurons, as well as the differentiation of GDNF-modified ADSCs was detected by immunofluorescence staining.RESULTS: GDNF appeared in ADSCs supematant at 24 hours after Ad-GDNF transfection and reached a peak at 72 hours.There was approximately 80% GFP-positive labeled in ADSCs. The tyrosinase hydroxylase staining results demonstrated that the rate of survival DN neurons were significantly increased than in DA neurons cultured alone, co-cultured group of GFP-modified ADSCs and GDNF-modified ADSCs groups (55%, 15%, 25%, P < 0.01). However, there were no co-expressing TH and GFP positive cells appeared at 7 days of co-culture, which indicated that the co-cultured condition was not available to ADSCs differentiation.CONCLUSION: The co-cultured of GDNF modified ADSCs and DN neurons can promote the survival and growth of cultured DN neurons, however, it can not induce ADSCs differentiate into DN neurons.

Behavioral changes in rats with focal cerebral ischemia after directional transplantation of bone marrow mesenchymal stem cells / 中国组织工程研究

BACKGROUND: Transplantation of bone marrow mesenchymal stem cells (MSCs) has been used in the field of repair of nerve injury. Brain stereotactic transplantation and transvascular transplantation are two transplantation methods. OBJECTIVE: We infused MSCs into rat peripheral cerebral infarct focus, in order to investigate the improvement of rat neurological dysfunction by forelimb use asymmetry test and postural reflex test.DESIGN: A randomized controlled animal experiment. SETTING: Department of Neurology, First Hospital of Jilin University.MATERIALS: This study was performed at the Key Laboratory of Zoonosis, Ministry of Education, Institute of Zoonosis, Jilin University between October 2006 and April 2007. Healthy male Wistar rats of clean grade, weighing 250-280 g, were provided by the Laboratory Animal Center of Jilin University. The protocol was performed in accordance with ethical guidelines for the use and care of animals.METHODS: MSCs from healthy adult volunteers were in vitro cultured and proliferated by density gradient separation and adherence screening method. Their immunophenotypes were identified by a flow cytometer. The Wistar rats were randomized into 5 groups with 10 rats in each: normal control group, sham-operated group, model group, serum-free DMEM-treated group (DMEM group) and MSCs -treated group (MSCs group). Rat models of cerebral ischemia/reperfusion were developed by occluding rat right middle cerebral artery following suture occlusion method modified by Longa et al. Rats in the normal control group were untouched. In the sham-operated group, operation was not ended till cervical interior and exterior arteries were exposed and sutured, and the other disposals were the same as those in the model group. At ischemia 90 minutes reperfusion 1 hour, a stereotaxic apparatus was used to take rat right peripheral cerebral ischemic region as transplantation site: 3 mm lateral to, 1mm caudal to and 4 mm posterior to Bregma. Rats in the MSCs group were slowly injected 5 μL BrdU-labeled MSCs (4×1011 L-1) serum-free medium. Rats in the DMEM group were injected 5 μL serum-free medium. After perfusion, inserted needle was retained for 5 minutes and then slowly withdrawn in order to avoid the back flow of liquid from needle pole. The survival of MSCs in rats was detected by immunohistochemical technique, and rat behavioral changes of observed on days1, 3, 7 and 28 after transplantation by forelimb use asymmetry test and postural reflex test.MAIN OUTCOME MEASURES: ① The immunophenotype of MSCs were identified by a flow cytometer. ② The survival of transplanted MSCs in the rat brain. ③ Rat behavioral changes. RESULTS: All the 50 rats were included in the final analysis. ① High purity of MSCs were harvested in the experiment. Flow cytometer detection showed that both CD44 and CD29 were positive, while CD34, CD45 and CD31 were negative. ② MSCs transplanted into the brains of rats in the MSCs group gathered in the peripheral cerebral ischemic region and survived. ③ Behavioral scores of rats in the MSCs group were significantly lower than those in the other groups (P < 0.05). They were gradually decreased with time after transplantation, and reached the valley value on day 7 after transplantation (P < 0.01). CONCLUSION: Neurological function of rats recovers in all the groups except normal control group. But the recovery differs in different groups, and neurological function of rats in the MSCs group recovers better than that in other groups.

Isolation and identification of hepatic stem cells from rat fetal liver in vitro / 吉林大学学报(医学版)

Objective To isolate and cultivate hepatic stem cells(HSCs) from rat fetal liver in vitro and identify their biological features.Methods Collagenase perfusion method and mechanical cutting method were used to isolate HSCs from rat fetal liver which were then cultivated by H-DMEM containing 10% fetal bovine serum.The cell surface antigen expression of HSCs was observed with immunocytochemical method under confocal laser scanning microscope.Results The isolated HSCs from rat fetal liver grew to monolayer 5 d after cultivation in vitro.They presented pykno-round cells and distinct borderline under the light microscope.After 8 d the cells grew like epithelium.These cells expressed AFP antigen,CK 18 and CK19.Conclusion The cultivated cells are proved to be HSCs and can proliferate quickly in vitro.

L-type calcium channels of human mesenchymal stem cells / 中国免疫学杂志

Objective:To investigate L-type calcium channels of human mesenchymal stem cells(hMSCs) cultured in vitro.Methods:hMSCs were isolated,cultured by Percoll gradient centrifugation,adherence to plastic flask and monoclonal culture.Immunophenotypes of hMSCs were detected by immunofluorescence and flow cytometry techniques.Whole cell patch-clamp technique was used to observe the change of L-type calcium channels of human mesenchymal stem cells(hMSCs).Results:High homogenous hMSCs had been isolated and cultured in vitro.hMSCs had unique immunophenotypes and they were positive for CD44,CD29,c-kit but negative for CD34,CD31,CD54.Calcium currents could be found in 40 percent of hMSCs,peak currents of calcium(I Ca-Peak ) were (102.67?19.06)pA from +20 mV to +30 mV.The currents could be inhibited by Cd~ 2+ (20 ?mol/L).Conclusion:Undifferentiated hMSCs express less L-type calcium channes,regulation of the chanels might contribute to directional differentiation.