Learning Molecular Representation in a Cell.

Predicting drug efficacy and safety in vivo requires information on biological responses (e.g., cell morphology and gene expression) to small molecule perturbations. However, current molecular representation learning methods do not provide a comprehensive view of cell states under these perturbations and struggle to remove noise, hindering model generalization. We introduce the Information Alignment (InfoAlign) approach to learn molecular representations through the information bottleneck method in cells. We integrate molecules and cellular response data as nodes into a context graph, connecting them with weighted edges based on chemical, biological, and computational criteria. For each molecule in a training batch, InfoAlign optimizes the encoder's latent representation with a minimality objective to discard redundant structural information. A sufficiency objective decodes the representation to align with different feature spaces from the molecule's neighborhood in the context graph. We demonstrate that the proposed sufficiency objective for alignment is tighter than existing encoder-based contrastive methods. Empirically, we validate representations from InfoAlign in two downstream tasks: molecular property prediction against up to 19 baseline methods across four datasets, plus zero-shot molecule-morphology matching.

Metabolic signatures and risk of sarcopenia in suburb-dwelling older individuals by LC-MS-based untargeted metabonomics.

Background: Untargeted metabonomics has provided new insight into the pathogenesis of sarcopenia. In this study, we explored plasma metabolic signatures linked to a heightened risk of sarcopenia in a cohort study by LC-MS-based untargeted metabonomics. Methods: In this nested case-control study from the Adult Physical Fitness and Health Cohort Study (APFHCS), we collected blood plasma samples from 30 new-onset sarcopenia subjects (mean age 73.2 ± 5.6 years) and 30 healthy controls (mean age 74.2 ± 4.6 years) matched by age, sex, BMI, lifestyle, and comorbidities. An untargeted metabolomics methodology was employed to discern the metabolomic profile alterations present in individuals exhibiting newly diagnosed sarcopenia. Results: In comparing individuals with new-onset sarcopenia to normal controls, a comprehensive analysis using liquid chromatography-mass spectrometry (LC-MS) identified a total of 62 metabolites, predominantly comprising lipids, lipid-like molecules, organic acids, and derivatives. Receiver operating characteristic (ROC) curve analysis indicated that the three metabolites hypoxanthine (AUC=0.819, 95% CI=0.711-0.927), L-2-amino-3-oxobutanoic acid (AUC=0.733, 95% CI=0.598-0.868) and PC(14:0/20:2(11Z,14Z)) (AUC= 0.717, 95% CI=0.587-0.846) had the highest areas under the curve. Then, these significant metabolites were observed to be notably enriched in four distinct metabolic pathways, namely, "purine metabolism"; "parathyroid hormone synthesis, secretion and action"; "choline metabolism in cancer"; and "tuberculosis". Conclusion: The current investigation elucidates the metabolic perturbations observed in individuals diagnosed with sarcopenia. The identified metabolites hold promise as potential biomarkers, offering avenues for exploring the underlying pathological mechanisms associated with sarcopenia.

Physical performance and muscle strength rather than muscle mass are predictor of all-cause mortality in hemodialysis patients.

Objectives: Patients undergoing maintenance hemodialysis usually suffer a high burden of poor functional status. The aim of this study was to investigate the association between muscle mass, muscle strength as well as physical performance with all-cause mortality in hemodialysis patients. Methods: 923 hemodialysis patients (565 men, mean aged 61.3 ± 12.7 years) were included from eight facilities in Tianjin and Shanghai of China from 2019 to 2021. Muscle mass was evaluated by skeletal muscle index (SMI) and muscle strength was assessed by handgrip strength. Different measures of physical performance were measured via gait speed, Timed Up and Go Test (TUGT) and short physical performance battery (SPPB). Cox proportional hazards regression models were used to determine the adjusted hazard ratios (HRs) of mortality with 95% confidence intervals (95% CIs) for baseline muscle mass, muscle strength and different measures of physical performance. Additionally, the area under the Receiver Operating Characteristic (ROC) curves were constructed to determine which index is a better predictor of mortality. Results: During a median follow-up of 14 (12-17 months), 79 (8.6%) patients died. Using the Cox regression analysis, we founded that muscle strength and physical performance rather than muscle mass were significantly negatively associated with mortality. The C-index for different measures of physical performance in predicting mortality were 0.709 for SPPB, 0.7 for TUGT and 0.678 for gait speed, respectively. The C-index for muscle strength was 0.635, and the ability of prediction was significantly lower than the physical performance. Conclusions: Physical performance seems to a better indicator of mortality than muscle mass and strength in hemodialysis patients. Simple measures of physical performance may be appropriately used as a screening tool targeting high-risk hemodialysis patients for the prevention of mortality.

Asiaticoside Prevents Oxidative Stress and Apoptosis in Endothelial Cells by Activating ROS-dependent p53/Bcl-2/Caspase-3 Signaling Pathway.

BACKGROUND: Asiaticoside (AC) is a triterpenoid saponin found in Centella asiatica (L.) urban extract that has a wide range of pharmacological properties. Our previous study demonstrated that AC could promote angiogenesis in diabetic wounds, but the specific mechanisms remain unknown. OBJECTIVE: This study aimed to examine the effectiveness and mechanism of AC on human umbilical vein endothelial cells (HUVECs) exposed to tert-butyl hydroperoxide (t-BHP) toxicity. METHODS: Senescence was confirmed using senescence-associated betagalactosidase (SA-ß-gal) activity and expression of the cell cycle phase markers p16 and p21. The levels of SOD, NO, MDA, GSH-Px, and ROS were tested. Furthermore, several cell death-related genes and proteins (p53, Bax, Bcl-2 and Caspase-3) were assessed with RT-qPCR and Western blotting. RESULTS: AC significantly reduced SA-ß-gal activity, with both the suppression of cellcycle inhibitors p16 and p21. We also found that the induced oxidative stress and apoptosis caused by t-BHP treatment resulted in the decrease of antioxidant enzymes activities, the surge of ROS and MDA, the up-regulation of p53, Bax and caspase-3, and the decrease of SOD, NO, GSH-Px and Bcl-2. These biochemical changes were all reversed by treatment with varying doses of AC. CONCLUSION: AC alleviates t-BHP-induced oxidative injury and apoptosis in HUVECs through the ROS-dependent p53/Bcl-2/Caspase-3 signaling pathway. It may be a potential antioxidant applied in metabolic disorders and pharmaceutical products.

The effects of the home-based exercise during COVID-19 school closure on the physical fitness of preschool children in China.

Background: Social distancing and school closures during the COVID-19 pandemic reduced the physical activities of the preschool children living in China. However, the effects of home-based exercise on the physical fitness of Chinese preschool children during COVID-19 school closures are still unknown. This study aimed to investigate the effects of home-based exercise on the physical fitness of Chinese preschool children during COVID-19 school closure. Methods: In this retrospective analysis, data from 1,608 Chinese preschool children (aged 3-5.5 years) in a second-tier city of Guangdong Province of China (Zhongshan city) were extracted from three successive National Physical Fitness Measurement (NPFM) from 2019 to 2021. NPFM consists of weight, height, and six subtests of physical fitness including 10-m shuttle run test (SRT), standing long jump (SLJ), balance beam walking (BBW), sit-and-reach (SR), tennis throwing (TT), and double-leg timed hop (DTH) tests. The change differences or change ratios of all the items in NPFM between any two successive years from 2019 to 2021 were compared. The exercise profiles about home-based and outdoor exercise before, during, and after COVID-19 school closure were obtained from 185 preschool children via retrospective telephone survey. Results: Between 2019 and 2021, 1,608 preschool children were included in this study. We observed larger changes in SLJ, SR, TT, and DTH tests during school closure than after school closure. But the children showed lower reduction rates in the completion time of SRT and BBW. During school closure, higher change ratios in SLJ and TT were observed in the children primarily participating in home-based exercise than those primarily participating in outdoor exercise. However, no statistical differences were observed in the changes in SRT and BBW between home-based and outdoor training groups. Conclusion: The home-based exercise program might be an alternative approach to improve the physical fitness of preschool children during COVID-19 school closure, but could not be beneficial to speed-agility and balance functions. A specific guideline geared toward a home-based exercise program during the COVID-19 outbreak is highly needed.

Sleep disturbances and sleep patterns in children with tic disorder: A case-control study.

Study objectives: To characterize sleep disturbances and sleep patterns in children with Tic disorder (TD), and explore their association with TD severity and types. Methods: A case-control study was conducted in 271 children with TD recruited from a clinical setting and 271 non-TD children recruited from a primary school, matched by age (mean = 8.47 years, SD = 1.53 years) and gender (15.1% female). The Children's Sleep Habits Questionnaire (CSHQ) was used to assess sleep patterns and sleep disturbances. The TD types and severity were assessed with the Yale Global Tic Severity Scale (YGTSS). Results: The TD children scored higher on CSHQ total score than non-TD group (t = 29.50, p < 0.001) and demonstrated severer global sleep disturbance. Compared to non-TD children, TD children presented with increased risks for global sleep disturbance (aOR: 1.95; 95% CI = 1.20-3.06), and most specific sleep disturbances, including bedtime resistance (aOR: 3.15; 95% CI = 1.96-5.06), sleep onset delay (aOR: 3.43; 95% CI = 1.58-7.46), sleep anxiety (aOR: 2.83; 95%CI = 1.83-4.38), parasomnias (aOR: 3.68; 95% CI = 2.02-6.62), night waking (aOR: 9.29; 95% CI = 2.64-32.65), sleep disordered breathing (aOR: 1.72; 95% CI = 1.03-2.90) and daytime sleepiness (aOR: 1.72; 95% CI = 1.09-2.74). Children with mild and moderate tics, Provisional Tic Disorder (PTD), Chronic Tic Disorder (CTD) and Tourette Syndrome (TS) presented with more global and more specific sleep disturbances. In addition, combined ADHD, etc. Conclusion: Children with TD are major risks for increased sleep disturbances, especially for those with severe and chronic symptoms. Furthermore, comorbid ADHD increases risk in certain areas of sleep. These findings highlight the importance to consider sleep outcomes in the assessment and treatment for children with TD.

Poor Physical Capacity Combined With High Body Fat Percentage as an Independent Risk Factor for Incident Hypertension in Chinese Suburb-Dwelling Older Adults.

Background: This study examined the effects of poor physical capacity and high body fat percentage (BF%) on the incidence of hypertension in Chinese suburb-dwelling older adults. Methods: This study was conducted on 368 Chinese suburb-dwelling participants aged ≥ 60 years without hypertension (mean age: 66.74 ± 5.59 years, 48.9% men). Poor physical capacity is defined by the Asian Working Group for Sarcopenia (AWGS) criteria as grip strength < 26 kg for men and < 18 kg for women or walking speed <0.8 m/s. High BF% was defined as values that are greater than the upper tertile for BF% as stratified by sex. The outcome was the incidence of hypertension. Results: Overall, 5.7% of subjects had both poor physical capacity and high BF%. After the average follow-up duration of 2 years, the incidence of hypertension was 39.7%, and those experiencing both poor physical capacity and high BF% had the highest incidence (81.0%). After multivariate adjustments, the incidence of hypertension was associated with the combination of poor physical capacity and high BF% [odds ratio (OR) = 6.43, 95% CI = 1.91-21.64] but not solely with poor physical capacity (OR = 1.11, 95% CI = 0.55-2.25) or only high BF% (OR = 1.37, 95% CI = 0.80-2.34). Conclusion: The combination of poor physical capacity and high BF% can significantly increase the incidence of hypertension in Chinese suburb-dwelling older adults. For hypertension prevention, ideally, we should strive toward decreasing body fat mass while simultaneously improving physical capacity.

Both Short and Long Sleep Durations are Risk Factors for Sarcopenia in Suburban-Dwelling Older Chinese Individuals: A 3-Year Longitudinal Study.

Background: Few longitudinal studies have explored exploring the relationship between sleep duration and sarcopenia. Evidence concerning the relationship between sleep duration and sarcopenia is limited and inconsistent. The purpose of this 3-year prospective study was to explore whether sleep duration was associated with sarcopenia onset in suburb-dwelling older Chinese individuals. Methods: This was a prospective study that included 754 Chinese suburb-dwelling men and women aged ≥60 years (men n=327, mean age 65.24± 4.87 years) who were not initially diagnosed with sarcopenia. We defined sarcopenia using the diagnostic algorithm recommended by the Asian Working Group for Sarcopenia. Self-reported sleep duration was a component of the interview measured by trained interviewers. Subjects were categorized into 3 groups at baseline [short: <6 h, medium: 6-8 h, and long: >8 h]. Results: The incidence of sarcopenia during the 3-year follow-up was 12.2%. Multivariate logistic regression analyses showed that after adjustments for potential confounders long sleep duration was independently associated with sarcopenia incidence from baseline through the 3-year follow-up: when using the 6-8 h sleep duration group as a reference, the adjusted ORs for sarcopenia of the groups who slept <6 and >8 hours were 2.74 (95% CI 1.05-7.13) and 1.84 (95% CI 1.07-3.14). Conclusion: Both short and long sleep durations were associated with a greater incidence of sarcopenia. Thus, sleep duration should be considered when developing prevention and management strategies for sarcopenia.

A Novel Perspective on the Proactive and Reactive Controls of Executive Function in Chronic Stroke Patients.

Objectives: To investigate the proactive and reactive control process when executing a complex task in patients with stroke. Proactive control is the preparatory process before the target stimulus, whereas reactive control is an imperative resolution of interference after the target stimulus. Methods: In total, 17 patients with chronic stroke and 17 healthy individuals were recruited. The proactive and reactive control of executive function was assessed by the task-switching paradigm and the AX version of the Continuous Performance Task (AX-CPT). The general executive function was assessed by Color Trial Test (CTT) and Stroop Test. The behavioral data of the task-switching paradigm were analyzed by a three-way repeated-measures ANOVA, and the AX-CPT data were analyzed by two-way repeated-measures ANOVA. Results: For efficiency scores in the task-switching paradigm, trial (repeat vs. switch) × group (stroke or control group) interaction effect was significant. Post-hoc analysis on trial × group effect showed a significant between-trial difference in accuracy rates in the repeat trial in the control group regardless of 100 or 50% validity. For the AX-CPT, the main effects of condition and group on response time were statistically significant. The interaction effect of condition (AY or BX) × group (stroke or control group) was also significant. Post-hoc analysis for condition × group indicated that the stroke group had a significantly longer response time in the BX condition than the control group and longer completion time in CTT2 and larger word interference for completion time in the Stroop test than the control cohort. Conclusions: Post-stroke survivors showed deficits in the performance of proactive control but not in the performance of reactive control. Deficits in proactive control may be related to the impairment of working memory. Interventions that focus on proactive control may result in improved clinical outcomes.

A Study on the Relationship between Postural Control and Pain-Related Clinical Outcomes in Patients with Chronic Nonspecific Low Back Pain.

OBJECTIVES: To explore the relationship between postural control and pain-related clinical outcomes in patients with chronic nonspecific low back pain (cNLBP). METHODS: Participants with cNLBP and healthy individuals were recruited. Muscle activities were recorded during internal and external perturbation tasks. Postural control capacity was assessed by muscle onset time and integrals of electromyography (iEMGs) of postural muscles during the phases of anticipatory postural adjustments (APAs) and compensatory postural adjustments (CPAs). Correlation analysis was employed to investigate the relationship between postural control capacity, pain, and disability. RESULTS: Twenty-seven patients with cNLBP and 27 healthy participants were recruited. Gastrocnemius (GA) muscle onset time was earlier in the cNLBP group than in the control group in the internal perturbation task. The onset time of GA and erector spinae (ES) of the cNLBP group was later than that of the controls in the external perturbation task. Disability level moderately correlated with the iEMGs of rectus abdominis (RA), GA, and external oblique (EO) during APAs. Pain score moderately correlated with the iEMGs of RA, EO, and ES during CPAs of perturbation tasks. CONCLUSION: cNLBP participants had altered muscle activation strategy to maintain postural stability in response to perturbation. This study further discovered that pain-related disabilities of cNLBP participants were likely related to the APAs capacity, whereas the pain intensity may relate to the CPAs capacity. Pain and disability may therefore be related to the control process of the posture-related muscles.

High-quality genome assembly of Metaphire vulgaris.

Earthworms enrich the soil and protect the health of their ecological environment. Previous studies on these invertebrates determined their protein content, hormone secretions, medicinal value, and ecological habits, but their whole genomic sequence remains incomplete. We performed whole genome sequencing of Metaphire vulgaris (Chen, 1930), which belongs to the genus Metaphire of the family Megascolecidae. The genome assembly was 729 Mb, with a N50 contig size of 4.2 Mb. In total, 559 contigs were anchored to 41 chromosomes according to the results of Hi-C (High-throughput Chromosome Conformation Capture) technology, which was confirmed by karyological analysis. A comparison of the genomic sequences and genes indicated that there was a whole-genome duplication in M. vulgaris followed by several chromosome fusion events. Hox genes and lumbrokinase genes were identified as partial clusters surrounding the genome. Our high-quality genome assembly of M. vulgaris will provide valuable information for gene function and evolutionary studies in earthworms.

The association between pelvic asymmetry and non-specific chronic low back pain as assessed by the global postural system.

BACKGROUND: Empirical evidence that demonstrates the relationship between pelvic asymmetry and non-specific chronic low back pain (NCLBP) is currently lacking. OBJECTIVE: To establish the reliability of the Global Postural System (GPS) in assessing pelvic asymmetry and identify the association between pelvic asymmetry parameters and the occurrence of NCLBP in young adults. DESIGN: A cross-sectional, regression study. METHODS: People who were aged between 18 and 30 and were diagnosed with NCLBP were recruited. Healthy individuals who were matched for age, sex, and education level were recruited as controls. Global Postural System (GPS) was employed to assess pelvic asymmetry. Prior to exploring the association, the reliability of GPS was assessed by the ICC (2, k) for interrater reliability, ICC (3, k) for intra-rater reliability, standard error and minimal detectable difference. Bivariate correlation analysis and logistic regression analysis were used to determine the relationship between pelvic asymmetry and the occurrence of NCLBP. RESULTS: Twenty-eight healthy participants and 28 people with NCLBP were recruited. Moderate to excellent ICCs were observed for the inter-rater and intra-rater reliability of most postural parameters. The bivariate correlation analysis indicated that age, body mass index and pelvic asymmetry parameters were related to the occurrence of NCLBP. Pelvic angle asymmetry (odds ratio = 1.17), and asymmetry of the distance between the posterior superior iliac spine and the floor (odds ratio = 1.21) were associated with NCLBP. LIMITATIONS: This study did not explore the causal relationship between pelvic asymmetry in the sagittal plane/pelvic asymmetry in the transverse plane and the occurrence of NCLBP. The interpretation of the results may not be generalized beyond the sample population. CONCLUSIONS: The GPS is a reliable method to assess pelvic asymmetry in a clinical setting. Two pelvic parameters were associated with the presence of NLBP. Measurement of pelvic asymmetry may assist in the early identification of potential occurrence of NCLBP but further work is required.

User testing of the psychometric properties of pictorial-based disability assessment Longshi Scale by healthcare professionals and non-professionals: a Chinese study in Shenzhen.

OBJECTIVE: The aim of this study was to validate a novel pictorial-based Longshi Scale for evaluating a patient's disability by healthcare professionals and non-professionals. DESIGN: Prospective study. SETTING: Rehabilitation departments from a grade A, class 3 public hospital, a grade B, class 2 public hospital, and a private hospital and seven community rehabilitation centers. SUBJECTS: A total of 618 patients and 251 patients with functional disabilities were recruited in a two-phase study, respectively. MAIN MEASURES: Outcome measure: pictorial scale of activities of daily living (ADLs, Longshi Scale). Reference measure: Barthel Index. The Spearman correlation coefficient was used to analyze the validity of Longshi Scale against Barthel Index. RESULTS: In phase 1 study, from March 2016 to August 2016, the results demonstrated that the Longshi Scale was both reliable and valid (intraclass correlation coefficient based on two-way random effect (ICC2,1) = 0.877-0.974 for intra-rater reliability; ICC2,1 = 0.928-0.979; κ = 0.679-1.000 for inter-rater reliability; intraclass correlation coefficient based on one-way random effect (ICC1,1) = 0.921-0.984 for test-retest reliability and Spearman correlation coefficient = 0.836-0.899). In the second phase, in March 2018, results further demonstrated that the Longshi Scale had good inter-rater and intra-rater reliability among healthcare professionals and non-professionals including therapists, interns, and personal care aids (ICC1,1 = 0.822-0.882 on Day 1; ICC1,1 = 0.842-0.899 on Day 7 for inter-rater reliability). In addition, the Longshi Scale decreased assessment time significantly, compared with the Barthel Index assessment (P < 0.01). CONCLUSION: The Longshi Scale could potentially provide an efficient way for healthcare professionals and non-professionals who may have minimal training to assess the ADLs of functionally disabled patients.

The Effect of Judo Training on Set-Shifting in School Children.

Improving executive functions (EFs) is desirable as they are considered to be critical for academic attainment and mental wellness in children. The aim of this study was to explore the effect of Judo training on the set-shifting function using a spatial task-switching paradigm. Protocol 1 compared the set-shifting ability of Judo players with age-matched healthy individuals. Protocol 2 compared the difference in EFs between children who underwent Judo training (intervention) and age-matched controls. EFs were assessed by a spatial task-switching test. Error rates and response times were analysed using two-way repeated-measures ANOVA. Protocol 1. The group effect on error rates was significant. The trial type × group effect was significant in the Judo group. Error rates in the Judo group were lower in the switch trials than the control group (p = 0.001). No significant group difference was seen in the repeat trials (p = 0.764). Protocol 2. The time × trial type × group effect was significant. Post hoc analysis showed significantly lower error rates by the intervention group on switch trials compared to the control group (p = 0.006). Regular Judo training may potentially be an option for improving EFs in schoolchildren or in populations with executive dysfunction.

[Study of the effects of integrin-linked kinase on proliferation and differentiation of fibroblast in hypertrophic scar].

OBJECTIVE: To study the role of integrin-linked kinase (ILK) on the proliferation and differentiation of human fibroblast in hypertrophic scar and its effect on the scar formation. METHODS: The human scar fibroblasts were isolated and cultured in vitro. The cells were divided into 4 groups. (1) control group: only contains DMEM; (2) jetPRIME group: DMEM with 200 microl jetPRIME buffer and 4 microl jetPRIME; (3) ILK siRNA group: DMEM and ILK siRNA; (4) ILK cDNA group: DMEM and ILK cDNA. The cell proliferation was detected by XTT assay and the mRNA and protein expressions of ILK and alpha-SMA were detected by Real-time qPCR and Western blot. RESULTS: (1) XTT results showed that the cellular proliferation level after 48 h in four groups were 0.820 +/- 0.065, 0.873 +/- 0.041, 0.554 +/- 0.013 and 1.296 +/- 0.094, respectively. The cellular proliferation curve showed that the cellular proliferation level was very flat in ILK siRNA group while the cellular proliferation level gradually increased from 12 h. 48 h after transfection, the cellular proliferation level in ILK siRNA group was significant lower than those in other groups (P value were 0.021, 0.034, 0), while the cellular proliferation level in ILK cDNA group was the highest among all 4 groups (P value were 0.017, 0.009, 0). (2) The Real-time qPCR showed that the expressions of ILK mRNA and alpha-SMA mRNA were 0.693 +/- 0.412 and 0.422 +/- 0.037 in control group, were 0.621 +/- 0.183 and 0.388 +/- 0.005 in jetPRIME group, were 0.052 +/- 0.019 and 0.073 +/- 0.023 in ILK siRNA group, were 240.193 +/- 35.170 and 138.056 +/- 24.060 in ILK cDNA group. The expressions of ILK mRNA and alpha-SMA mRNA in ILK siRNA group were significantly lower than those in other three groups (P < 0.05). And the expressions of ILK mRNA and alpha-SMA mRNA in ILK cDNA group were significantly higher than those in other three groups (P < 0.05). (3) The Western blot also showed that the expression of ILK and alpha-SMA proteins were decreased in ILK siRNA group and increased in ILK cDNA group. CONCLUSION: ILK may promote the proliferation and differentiation of human scar fibroblast. It may play an important role in scar formation and contracture.

[Expression of integrin-linked kinase in fibroblasts of scar induced by cobalt chloride and its effect on cell proliferation].

OBJECTIVE: To explore the expression of integrin-linked kinase (ILK) in fibroblasts (Fbs) of scar induced by cobalt chloride (CoCl2) and its effect on cell proliferation. METHODS: The human hypertrophic scar Fbs of seven patients were isolated and cultured in vitro. Cells from the 5th to the 6th passages were used in the experiment. Six bottles of Fbs were obtained from each of the seven patients, and they were respectively cultured with DMEM nutrient solution containing CoCl2 in the concentration of 0, 50, 100, 150, 200, and 250 µmol/L for 24 h. The expression of ILK mRNA was determined with real-time fluorescence quantitative PCR. Fbs were stimulated by CoCl2 in the most suitable concentration (100 µmol/L) and the protein expression of ILK was determined 0, 1, 2, 4, 12, and 24 h after the stimulation. Then the Fbs were divided into control group (cultured with nutrient solution), negative control group (transfected with con-siRNA), and ILK siRNA group (transfected with ILK siRNA). They were cultured with nutrient solution containing CoCl2 in different concentrations 24 h after transfection, with 4 wells for each concentration in each group. The cell proliferation was detected by XTT assay. Data were processed with one-way analysis of variance (ANOVA) and ANOVA for repeated measurement, and LSD method was used in multiple comparisons. RESULTS: The expression level of ILK mRNA was highest in Fbs cultured with 100 µmol/L CoCl2 for 24 h, with significant difference compared with those of Fbs cultured with other concentrations of CoCl2 (F = 50.958, P < 0.001). The expression of ILK protein in Fbs cultured with 100 µmol/L CoCl2 for 1 h (0.243 ± 0.009) was lower than that cultured for 0 h (0.387 ± 0.017), and it started to increase from 2 h (0.361 ± 0.010), and exaggerated at 4 h (0.584 ± 0.028), 12 h (0.730 ± 0.029), and 24 h (0.785 ± 0.031). The expression levels of ILK protein at 1, 4, 12, 24 h were statistically different from that at 0 h (P values all below 0.05). XTT showed that cell proliferation level was highest in control group when cultured with 100 µmol/L CoCl2 (F = 488.026, P < 0.001), which decreased from 150 µmol/L. The cell proliferation level in control group cultured with 250 µmol/L CoCl2 was significantly lower than that with 0 µmol/L (P values all below 0.05). There was no significant change in cell proliferation in ILK siRNA group among different concentrations of CoCl2 (F = 2.542, P = 0.056). The cell proliferation level in ILK siRNA group was significantly lower than that in control group and negative control group (F = 2519.542, P < 0.001). CONCLUSIONS: ILK may be a key protein in response of hypoxia in Fbs. The mild hypoxia can stimulate the expression of ILK and promote the proliferation of Fbs, while severe hypoxia can reduce the expression of ILK and inhibit cell proliferation.

[The effects of intergrin-linked kinase on angiogenesis in hypertrophic scar].

OBJECTIVE: To investigate the effects and regulatory mechanism of ILK on angiogenesis in hypertrophic scar. METHODS: The human scar microvascular endothelial cells (HSMECs) were isolated from 6 patients' hypertrophic scar in vitro. The HSMECs with good condition in 2nd to 4th generation were selected as experimental objectives. (1) HSMECs were divided into the blank control group (treated with routine culture), negative control group (treated with only Lipofectamine 2000), LY294002 group (incubated with 50 nmol/L LY294002), ILK siRNA group (incubated with 20 nmol/L ILK siRNA). RT-PCR and Western Blot were used to detect the expression of ILK mRNA and its protein after transfecion for 48 h. (2) The digested HSMECs of four groups were resuspended with DMEM without serum and then seeded onto the upper compartment of transwell insert which contained complete medium in its lower compartment. The cell migration experiment was stopped in 10 h and then the migrated cells were counted to analyze the effects of different interventions on the migration ability of HSMECs. (3) The thawed ECMatrix was put into each well of pre-colled 48-well tissue culture plate, and then the plate was put into the incubator at 37 degrees C to make it to become gel. The HSMECs of four groups were seeded onto the surface of the ECMatrix gel and were put into incubator. Eight random view-fields per well should be valued by the sheet of pattern recognition about angiogenesis after 8 hours to evaluate the ability of angiogenesis in vitro between four groups. RESULTS: (1) The expression of ILK mRNA (ILK mRNA = 0.829 +/- 0.109, t = 13.151, P = 0.006) and protein (ILK protein = 0.096 +/- 0.049, t = 36.656, P = 0.000) were both inhibited obviously in ILK siRNA group compared with the blank control group (ILK mRNA = 0.829 +/- 0.109, ILK protein = 1). And, the expression of ILK in LY294002 group was slightly lower than that of black control group, but there was no statistical difference. (2) The number of migrated cells in ILK siRNA group (88.111 +/- 3.079) and LY294002 group (138. 667 +/- 2.404) were respectively lower than that in blank control group (322.333 +/- 3.712, P < 0. 05) in 10th hour. (3) Compared to blank control group (4.333 +/- 0.191), the ability of angiogenesis in vitro decreased significantly ILK siRNA group (2.625 +/- 0.125) and LY294002 group (3.125 +/- 0.250), in which, the vascular network structures were not formed perfectly in 8th hour (P < 0.05). CONCLUSIONS: The ability of HSMECs' migration and angiogenesis in vitro are inhibited significantly when the expression of ILK is down-regulated. It reveals that ILK may play an role in the regulation of scar angiogenesis.

[Expression of integrin-linked kinase in human hypertrophic scar and its relationship with angiogenesis].

OBJECTIVE: To explore the expression of integrin-linked kinase (ILK) in scar in different growth stages, as well as its relationship with angiogenesis. METHODS: (1) Fifteen burn patients with scar formation time shorter than 6 months, ranging from 6 to 12 months, and longer than 12 months were hospitalized from December 2009 to December 2010. They were divided into A, B, and C groups according to the scar formation time, with 5 patients in each group. Scar specimens were harvested for observation of ILK expression with immunohistochemistry method, and ILK mRNA expression with real time fluorescence quantitative RT-PCR. (2) Microvascular endothelial cells (MEC) were isolated from scar tissue in A group and cultured in vitro, and then they were purified by immunomagnetic beads and identified with coagulation factor VIII marked by immunofluorescence (fibroblasts from human normal skin were used as control). The cultured cells in logarithmic growth phase were divided into control group (cultured with M131 medium containing microvascular growth supplement), transfection 1 group (transfected with empty plasmid), and transfection 2 group (transfected with ILK cDNA plasmid) according to the random number table. After 24 hours, the expressions of ILK mRNA, Flt-1 mRNA, and KDR mRNA were determined with real time fluorescence quantitative RT-PCR. Data were processed with one-way analysis of variance. RESULTS: Immunohistochemical observation showed that ILK in A group mainly expressed in the basal layer cells of epidermis, cytoplasm of fibroblasts, and MEC in scar, while ILK in B group only distributed in the basal layer cells of epidermis, but ILK expression in C group was not obvious. The expression of ILK mRNA in A group (0.34 ± 0.16) was significantly higher than those in B and C groups (0.17 ± 0.06, 0.07 ± 0.13, F = 37.007, P = 0.000). MEC grew up showing cobble stone formation after purification. The expression of coagulation factor VIII was positive in cytoplasm of purified MEC, while that was negative in fibroblast of human normal skin. The expressions of ILK mRNA (57.807 ± 5.556), KDR mRNA (0.836 ± 0.014), and Flt-1 mRNA (0.162 ± 0.005) in transfection 2 group were higher than those in control and transfection 1 groups (0.018 ± 0.003, 0.028 ± 0.020, 0.023 ± 0.004 and 0.042 ± 0.005, 0.039 ± 0.007, 0.046 ± 0.003; F(ILK) = 87.110, F(KDR) = 11.241, F(Flt) = 18.199, with P values all below 0.01). CONCLUSIONS: ILK mainly expressed in scar tissue with formation time shorter than 6 months, and it may affect vascularization of scar by regulating gene expressions of KDR and Flt-1 in MEC, which plays an important role in early scar formation.