Exosomes derived from induced cardiopulmonary progenitor cells alleviate acute lung injury in mice.

Cardiopulmonary progenitor cells (CPPs) constitute a minor subpopulation of cells that are commonly associated with heart and lung morphogenesis during embryonic development but completely subside after birth. This fact offers the possibility for the treatment of pulmonary heart disease (PHD), in which the lung and heart are both damaged. A reliable source of CPPs is urgently needed. In this study, we reprogrammed human cardiac fibroblasts (HCFs) into CPP-like cells (or induced CPPs, iCPPs) and evaluated the therapeutic potential of iCPP-derived exosomes for acute lung injury (ALI). iCPPs were created in passage 3 primary HCFs by overexpressing GLI1, WNT2, ISL1 and TBX5 (GWIT). Exosomes were isolated from the culture medium of passage 6-8 GWIT-iCPPs. A mouse ALI model was established by intratracheal instillation of LPS. Four hours after LPS instillation, ALI mice were treated with GWIT-iCPP-derived exosomes (5 × 109, 5 × 1010 particles/mL) via intratracheal instillation. We showed that GWIT-iCPPs could differentiate into cell lineages, such as cardiomyocyte-like cells, endothelial cells, smooth muscle cells and alveolar epithelial cells, in vitro. Transcription analysis revealed that GWIT-iCPPs have potential for heart and lung development. Intratracheal instillation of iCPP-derived exosomes dose-dependently alleviated LPS-induced ALI in mice by attenuating lung inflammation, promoting endothelial function and restoring capillary endothelial cells and the epithelial cells barrier. This study provides a potential new method for the prevention and treatment of cardiopulmonary injury, especially lung injury, and provides a new cell model for drug screening.

Cardiopulmonary progenitors facilitate cardiac repair via exosomal transfer of miR-27b-3p targeting the SIK1-CREB1 axis.

Ischemic heart disease, especially myocardial infarction (MI), is one of the leading causes of death worldwide, and desperately needs effective treatments, such as cell therapy. Cardiopulmonary progenitors (CPPs) are stem cells for both heart and lung, but their repairing role in damaged heart is still unknown. Here, we obtained CPPs from E9.5 mouse embryos, maintained their stemness while expanding, and identified their characteristics by scRNA-seq, flow cytometry, quantitative reverse transcription-polymerase chain reaction, and differentiation assays. Moreover, we employed mouse MI model to investigate whether CPPs could repair the injured heart. Our data identified that CPPs exhibit hybrid fibroblastic, endothelial, and mesenchymal state, and they could differentiate into cell lineages within the cardiopulmonary system. Moreover, intramyocardial injection of CPPs improves cardiac function through CPPs exosomes (CPPs-Exo) by promotion of cardiomyocytic proliferation and vascularization. To uncover the underlying mechanism, we used miRNA-seq, bulk RNA-seq, and bioinformatic approaches, and found the highly expressed miR-27b-3p in CPPs-Exo and its target gene Sik1, which can influence the transcriptional activity of CREB1. Therefore, we postulate that CPPs facilitate cardiac repair partially through the SIK1-CREB1 axis via exosomal miR-27b-3p. Our study offers a novel insight into the role of CPPs-Exo in heart repair and highlights the potential of CPPs-Exo as a promising therapeutic strategy for MI.

LDLR is an entry receptor for Crimean-Congo hemorrhagic fever virus.

Crimean-Congo hemorrhagic fever virus (CCHFV) is the most widespread tick-born zoonotic bunyavirus that causes severe hemorrhagic fever and death in humans. CCHFV enters the cell via clathrin-mediated endocytosis which is dependent on its surface glycoproteins. However, the cellular receptors that are required for CCHFV entry are unknown. Here we show that the low density lipoprotein receptor (LDLR) is an entry receptor for CCHFV. Genetic knockout of LDLR impairs viral infection in various CCHFV-susceptible human, monkey and mouse cells, which is restored upon reconstitution with ectopically-expressed LDLR. Mutagenesis studies indicate that the ligand binding domain (LBD) of LDLR is necessary for CCHFV infection. LDLR binds directly to CCHFV glycoprotein Gc with high affinity, which supports virus attachment and internalization into host cells. Consistently, a soluble sLDLR-Fc fusion protein or anti-LDLR blocking antibodies impair CCHFV infection into various susceptible cells. Furthermore, genetic knockout of LDLR or administration of an LDLR blocking antibody significantly reduces viral loads, pathological effects and death following CCHFV infection in mice. Our findings suggest that LDLR is an entry receptor for CCHFV and pharmacological targeting of LDLR may provide a strategy to prevent and treat Crimean-Congo hemorrhagic fever.

[Effect of cotton-moxibustion on chronic eczema and quality of life].

OBJECTIVE: To compare the therapeutic effect between cotton-moxibustion and compound flumetasone ointment, and observe the effect on quality of life in patients with chronic eczema. METHODS: A total of 66 patients with chronic eczema were randomized into an observation group (33 cases, 2 cases dropped off) and a control group (33 cases, 2 cases dropped off). In the observation group, cotton-moxibustion was adopted on target skin lesion, once a day, 3 cones a time. In the control group, external application of compound flumetasone ointment was given twice a day. The treatment for 3 weeks was required in the both groups. Before treatment and 1,2,3 weeks into treatment, scores of visual analogue scale (VAS), eczema area and severity index (EASI) and dermatology life quality index (DLQI) were observed, and the recurrence rate was evaluated in the follow-up one month after treatment. RESULTS: Compared before treatment, the VAS scores of 1,2,3 weeks into treatment, the EASI and DLQI scores of 2,3 weeks into treatment were decreased in the both groups (P<0.05), and those in the observation group were lower than the control group (P<0.05). The follow-up recurrence rate in the observation group were lower than the control group (P<0.05). CONCLUSION: Cotton-moxibustion can effectively improve the pruritus symptom, skin lesion and quality of life in the patients with chronic eczema, the therapeutic effect is superior to the external application of compound flumetasone ointment.

Monoterpenoid indole alkaloids from the stems of Kopsia officinalis.

Five new indole alkaloids, kopsiofficines H-L (1-5), along with fourteen known alkaloids (6-19) were isolated from the stems of Kopsia officinalis. Their structures were elucidated by extensive NMR, mass spectroscopic analyses and comparison to the reported data. All the isolated compounds were evaluated their anti-inflammatory activities by inhibiting IL-1ß, PGE2 and TNF-α secretion in lipopolysaccharide (LPS)-activated RAW264.7 cells. Compounds 2, 3, 6, 7, 11, 12, 15, and 17 show significant anti-inflammatory activities. These results demonstrate pharmacodynamic substance basis of these folkloric claims.

[Epidemiological study of clinical characteristics of Chinese Han ethnic women with polycystic ovary syndrome].

OBJECTIVE: To carry out an epidemiological study of clinical characteristics of Chinese Han ethnic women with polycystic ovary syndrome (PCOS). METHODS: According to Revised 2003 European Society for Human Reproduction and Embryology/American Society for Reproductive Medicine criteria, PCOS can be diagnosed with 2 manifestations out of oligo-or anovulation, clinical and/or biochemical signs of hyperandrogenism exclusion of other etiologies and polycystic ovaries. One thousand and twenty-seven women in reproductive age from one area in Jinan city were investigated and the clinical, metabolic characteristics of the PCOS patients were analyzed. RESULTS: (1) A total of 828 questionnaires were collected from 1027 women; the response rate was 80.62%. Eighty-five PCOS patients were diagnosed; PCOS accounted for 97.65% (83/85) in

[Effects of gonadotropin on the expression of growth differentiation factor 9 and 9B in mouse ovary].

OBJECTIVES: To study the effects of gonadotropin on the expression of growth differentiation factor-9 (GDF-9) and -9B (GDF-9B) in mouse ovary. METHODS: We chose follicles of mature mice cultured in vitro and mature BALB/c mice as our animal models. (1) In vivo experiment. Twenty mice were divided into two groups (groups A and B) randomly with ten mice in each group. Each mouse was injected with 10 IU pregnant mare's serum gonadotropin in group A and saline of the same volume was given to the other group. Twenty-four hours later we obtained ovarian tissue and immunohistochemistry and in situ hybridization were performed to detect the expression of GDF-9 and GDF-9B. (2) In vitro experiment. Fifty-eight separated follicles were divided into two groups (groups C and D) randomly and cultured in vitro. Thirty-two follicles in group C were cultured in medium with follicular stimulating hormone (FSH) for 72 hours while 26 were cultured without FSH. Immunohistochemistry was performed to detect the expression of GDF-9 in both groups. RESULTS: We abserved 370 follicles by immunohistochemistry. The weak positive (+ or ++) rate was 22.0% and 46.2% while the strong positive (+++ or ++++) rate was 22.6% and 9.1% in 186 follicles of group A. The weak positive (+ or ++) rate in group B was 42.4% and 42.9% while the strong positive (+++ or ++++) rate was 14.1% and 0.5%. The expression of GDF-9 was higher in group A than that of group B. The weak positive rate, positive rate and strong positive rate in group C was 28.1%, 53.1% and 18.8% while that in group D was 61.5%, 30.8% and 7.7%, respectively. The expression of GDF-9 was higher in group C than that of group D. We observed 362 follicles by in situ hybridization. The weak positive (+ or ++) rate was 31.5% and 46.2% while the strong positive (+++ or ++++) rate was 21.2% and 1.1% in 184 follicles of group A. The weak positive (+ or ++) rate in group B was 39.9% and 41.6% while the strong positive (+++ or ++++) rate was 15.7% and 2.8%. There were no differences in the expression of GDF-9B between groups A and B. CONCLUSION: Gonadotropin increases the expression of GDF-9 in vivo while FSH increases GDF-9 in vitro. Gonadotropin has no effects on the expression of GDF-9B.

[Study on the (TAAAA)n repeat polymorphism in sex hormone-binding globulin gene and the SHBG serum levels in putative association with the glucose metabolic status of Chinese patients suffering from polycystic ovarian syndrome in Shandong province].

OBJECTIVE: To investigate the relationship of (TAAAA)n repeat polymorphism in the promoter of the sex hormone-binding globulin (SHBG) gene and SHBG serum levels to the glucose metabolic status of Chinese polycystic ovary syndrome (PCOS) patients in Shandong province. METHODS: GeneScan method was used to detect and identify (TAAAA)n repeat number (alleles) and genotypes for 156 controls and 157 patients who were divided into normal glucose tolerance without hyperinsulinemia (NIR group) and with hyperinsulinemia (HI group) and abnormal glucose metabolic (AGM) group according to the results of oral glucose test and insulin resistant test; IRMA was used to measure serum SHBG for part of them. RESULTS: Five alleles containing (TAAAA) 6-10 repeats and 14 genotypes including 6/6, 6/7, 6/8, 6/9, 6/10, 7/7, 7/8, 7/9, 7/10, 8/8, 8/9, 8/10, 9/9, 9/10 repeats genotypes were present in the subjects. Genotype distribution of 6/10 repeats genotype is lower in PCOS than that in control, and 8/9 repeats genotype vice versa (P < 0.01); among PCOS subgroups, the eight repeat genotypes in NIR group is more frequent than that in HI group (P < 0.01), and 7/9 genotype distribution in AGM group is higher than that in NIR group and HI group(P < 0.05-0.01). The serum SHBG levels in homozygous genotype groups exhibit a sequence of 8/8 > 9/9 > 6/6, 7/7 repeats and the fall of serum SHBG trend is in reversed relation with the increase in body mass index (BMI), Homa-IR, and blood pressure. Serum SHBG levels in AGM exhibit a sequence of HI group < NIR group < control but show no statistical difference between both groups. CONCLUSION: This study reveals that the repeat number, alleles, genotypes and their distributions in Chinese women are very different from these in foreigners. Some special genotypes and low serum SHBG levels may be associated with PCOS and its glucose metabolic status; some special genotypes may influence Chinese serum SHBG and need more studies, but both SHBG gene polymorphism genotype and serum SHBG are not good indicators to find out the PCOS individual at high risk.

[Correlation between 4G and 5G genetypes distribution of plasminogen activator inhibitor-1 gene polymorphism in its promoter region with polycystic ovarian syndrome].

OBJECTIVE: To investigate the correlation of 4G and 5G genetypes distribution of plasminogen activator inhibitor-1 (PAI-1) gene polymorphism in its promoter region with polycystic ovary syndrome (PCOS). METHODS: In 101 Chinese PCOS patients and 42 women as control, 4G and 5G polymorphisms of PAI-1 gene were detected with PCR-restriction fragment length polymorphism (RFLP). Pregnant history, body mass index (BMI), waist-hip ratio (WHR) were collected and Homastasis Model Assessment score for insulin resistant (Homa-IR) and insulin sensitive index (ISI) in PCOS groups were calculated. RESULTS: The distributions of PAI-1 gene polymorphisms 4G type (4G/4G genetype) and 5G type (5G/5G, 4G/5G genetype) were different between the PCOS group and the control. The PCOS group had higher 4G type 57% (58/101) distribution than that of the control group 38% (16/42); 5G type is vise verse (P < 0.05). PCOS patients were divided into obese and non-obese sub-groups according to BMI. There were significant lower Homa-IR and higher ISI in non-obese subgroup than that in obese group (P < 0.05; P < 0.01). 68% (36/53) 4G type distribution in the non-obese is higher than that [48% (23/48)] in obese sub-group (P < 0.05). There were higher 4G type distribution 79% (11/14) and lower 5G type distribution 21% (3/14) in spontaneous miscarriage group than that of in non-spontaneous miscarriage group 38% (5/13) and 62% (8/13) among patients who impregnated at least once (n = 27, P < 0.05). CONCLUSION: PAI-1 gene polymorphism 4G genetype may be correlated with PCOS in Chinese women, especially in PCOS patients with non-obese PCOS and spontaneous miscarriage.

[In vitro maturation and fertilization of oocytes from unstimulated cycles in women with infertility due to polycystic ovary syndrome].

OBJECTIVE: To investigate the effect of in vitro maturation (IVM) and fertilization of oocytes from unstimulated cycles in women with infertility due to polycystic ovary syndrome (PCOS). METHODS: Seventy women with PCOS, who came to our clinic during March to were involved in this trial. June 2003. Every cycle was given human chorionic gonadotropin 10,000 IU 36 h before oocyte retrieval. Immature oocytes were matured in vitro and fertilized, and the resulting embryos were replaced back to the uterus. RESULTS: A total of 94 IVM cycles were performed and 1283 oocytes were obtained. The overall maturation, fertilization and cleavage rates were 65.3%, 66.0% and 48.0% respectively. Maturation, fertilization, cleavage and high quality embryo rates had no difference between metaphase I stage oocytes (69.7%, 71.7%, 52.2% and 26.1% respectively) and germinal vesical (GV) stage oocytes (67.7%, 66.4%, 47.6% and 24.1% respectively), but the rates in those oocytes which could not be classified (44.8%, 53.8%, 46.2% and 16.9% respectively) were much lower than the former two groups. After embryo transfer, 18 pregnancies were reported (24%). CONCLUSION: IVM/IVF-ET in unstimulated cycles is a feasible treatment for women with PCOS.

[Treatment of polycystic ovarian syndrome with anovulatory infertility by ultrasound-guided immature follicle aspiration].

OBJECTIVE: To investigate the effect of ultrasound-guided immature follicle aspiration (IMFA) on polycystic ovarian syndrome (PCOS) in women with anovulatory infertility, such as the endocrinology of the patients and the basic follicles number in the ovaries, and to observe the ovarian response to human menopausal gonadotropin (hMG) stimulation and pregnancy after the therapy. METHODS: Seventy-one PCOS patients were involved in this trial, and divided into 2 groups. Thirty-seven patients of group I were primed with slight amount of hMG, the other 34 patients did not use hMG. Ultrasound-guided IMFA was performed 36 hour after human chorionic gonadotropin (hCG) administration. In the next cycle, the basal endocrinology and the basic number of follicles were checked, then IMFA were performed continuously until the basic number of follicles were under 10 per ovary. Afterwards, hMG were used to stimulate ovulation and the pregnant results were followed up. RESULTS: There were 37 cases (88 cycles) of PCOS patients in group I and 34 cases (87 cycles) in group II. After 2 to 3 treatment cycles, the testosterone level became normal in all of patients. The basic follicle number decreased to less than 10 per ovary in 33 cases (89%) in group I and 28 cases (82%) in group II. After hCG stimulation, all of them ovulated. Only 2 patients developed slight ovarian hyperstimulation syndrome (OHSS). Within 3 months after the procedure, 36 patients (51%) became pregnant by controlled ovarian hyperstimulation (COH) protocol after IMFA. CONCLUSIONS: Immature follicle aspiration treatment can improve the abnormal endocrinology, decrease the basic follicle number of the ovary, and achieve pregnancy in following COH cycles, meanwhile, avoid OHSS in PCOS patients.

[Correlation between single nucleotide polymorphism of insulin receptor gene with polycystic ovary syndrome].

OBJECTIVE: To investigate the correlation of C/T single nucleotide polymorphism at exon 17 in tyrosine kinase domain of insulin receptor gene with polycystic ovary syndrome (PCOS). METHODS: One hundred and twenty patients with PCOS (PCOS group) and 40 normal females (control group) were included and the 1058 site single nucleotide polymorphism at exon 17 of insulin receptor gene (INSR) was detected with polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). The baseline parameters including levels of serum follicular stimulating hormone (FSH), luteinizing hormone (LH), prolactin (PRL), estradiol (E(2)), testosterone (T) and lipid, as well as the body mass index (BMI) and waist/hip ratio (WHR) were measured. Glucose tolerance and insulin releasing before and after loading with 75 g of glucose were also assayed. RESULTS: (1) BMI (25 +/- 4 vs. 23 +/- 3, P < 0.01), WHR (0.82 +/- 0.06 vs. 0.79 +/- 0.07, P < 0.05), LH [(9.2 +/- 6.1) U/L vs. (4.4 +/- 1.6) U/L, P < 0.01], T [(1.9 +/- 0.9) nmol/L vs. (1.0 +/- 0.6) nmol/L, P < 0.01] and E(2) [(205 +/- 119) pmol/L vs. (143 +/- 75) pmol/L, P < 0.01] of PCOS group were higher than those of control group. (2) The blood sugar and insulin levels of fasting, 30 min, 60 min, 120 min after glucose loading in obese PCOS (BMI >/= 25 kg/m(2)) were significantly higher than those in the non-obese PCOS (BMI < 25 kg/m(2)). (3) C/T single nucleotide polymorphism frequency at exon 17 of INSR in patients with PCOS was significantly higher than that in normal female (41% vs. 12.5%, P < 0.01). There was a significant difference between the frequency in non-obese PCOS (52.2%) and obese PCOS (25.5%, P < 0.01). (4) BMI in patients with T allele 23 +/- 3 of INSR single nucleotide polymorphism was lower than that with C allele (26 +/- 4, P < 0.05). CONCLUSIONS: (1) The body weight and gene single nucleotide polymorphism of INSR may play an important role in the occurrence of insulin resistance in patients with PCOS. (2) The 1058 site nucleotide polymorphism of insulin receptor gene is one of the susceptibility genes in patients with PCOS, especially in non-obese PCOS patients.

[Clinical analysis of multiple pregnancy reduction].

OBJECTIVE: To analyse the effect of the reduction of multiple pregnancy through transvaginal ultrasonic monitoring on the pregnancy outcome. METHODS: Eighty-four cases were divided into two groups according to whether they had vaginal hemorrhage before operation. And the pregnancy outcomes were analyzed. RESULTS: The abortion rate and preterm birth rate of the vaginal hemorrhage group were higher, and the difference was statistically significant. CONCLUSIONS: The reduction of multiple pregnancy through transvaginal ultrasonic monitoring is a safe operative method. But it is only a remedial treatment for multiple pregnancy, and how to prevent multiple pregnancy is of more practical value.

Comparison of primary breast cancer size by mammography and sonography.

PURPOSE: To compare tumor size by mammography and sonography and align with pathological results in primary breast cancer cases. MATERIALS AND METHODS: We retrospectively reviewed 95 primary breast cancer patients who underwent mammography and sonography from January 2011 to June 2012. The largest tumor diameter was chosen as sizing reference for each imaging modality. The measurements of mammography and sonography were considered concordant if they were within the measurement of pathological results±0.5 cm. Pearson's correlation coefficient was calculated for imaging results. RESULTS: The range of the maximum diameter was 0.6 cm-10.5 cm and mean value was 3.81±2.04 cm by pathological results, 0.7 cm-12.4 cm and 3.99±2.19 cm by mammography, and 0.9 cm-11.0 cm and 3.63±2.01 cm by sonography, respectively. Sonography (R: 0.754), underestimated tumor size, but had a better correlation with pathological tumor size compared to mammography (R: 0.676), which overestimated tumor size. CONCLUSIONS: Sonography is superior to mammography in assessment of primary breast cancer.

Electricity generation and brewery wastewater treatment from sequential anode-cathode microbial fuel cell.

A sequential anode-cathode double-chamber microbial fuel cell (MFC), in which the effluent of anode chamber was used as a continuous feed for an aerated cathode chamber, was constructed in this experiment to investigate the performance of brewery wastewater treatment in conjugation with electricity generation. Carbon fiber was used as anode and plain carbon felt with biofilm as cathode. When hydraulic retention time (HRT) was 14.7 h, a relatively high chemical oxygen demand (COD) removal efficiency of 91.7%-95.7% was achieved under long-term stable operation. The MFC displayed an open circuit voltage of 0.434 V and a maximum power density of 830 mW/m(3) at an external resistance of 300 Omega. To estimate the electrochemical performance of the MFC, electrochemical measurements were carried out and showed that polarization resistance of anode was the major limiting factor in the MFC. Since a high COD removal efficiency was achieved, we conclude that the sequential anode-cathode MFC constructed with bio-cathode in this experiment could provide a new approach for brewery wastewater treatment.

Effects of lead on 4-aminobiphenyl pharmacokinetics in liver, kidney, spleen, testes, heart, lung and hemoglobin adduct for rat model.

Lead affects almost every system in the body and 4-aminobiphenyl increases the incidence of bladder cancer among human exposed to aromatic amines, but little attention has been paid to the effects of lead (heavy metals) on the distribution and metabolic kinetics of 4-ABP (organic pollutants) in the organisms of the terrestrial ecosystems. In this study, male S.D. rats as model animals drinking tap water with and without lead with i.p. injection of 4-aminophenyl were used to study 4-aminobiphenyl pharmacokinetics with statistical analysis in three types of information systems: (1) hemoglobin adduct in the blood; (2) distribution concentrations in liver, kidney, spleen, testes, heart and lung; (3) relative weights of the six organs. Kinetic equations of 4-aminobiphenyl-hemoglobin adduct for two groups of rats drinking water with and without lead were all linear. Principal components were obtained based on three types of variants: (1) variants of distribution concentrations; (2) variants of relative weights; (3) variants of hemoglobin adduct, distribution concentrations and relative weights. Through a comparison of two groups of principal components, the result implied that lead changed 4-aminobiphenyl distribution kinetics in the six organs, had significant effects on the six organ relative weights, and had also significant effects on all thirteen variants as a whole. Correlation analysis of the principal components showed that lead could not significantly change the relation of hemoglobin adduct with time after dosing 4-aminobiphenyl. However, another result indicated that lead considerably improved the correlation between hemoglobin adduct and the thirteen variants as a whole. This implied that hemoglobin adduct could characterize all the thirteen variants as an index of 4-aminobiphenyl pharmacokinetics for the rats drinking water with lead, which conclusion was not suitable for the rats drinking water without lead. The research indicated that heavy metals existing in the organisms play an important role in the studies on pharmacotoxicology of organic pollutants. Frequently, various xenobiotics (heavy metals and organic pollutants) enter organisms simultaneously, therefore heavy metals should be considered comprehensively in the pharmacotoxicology of organic pollutants in animals in the terrestrial ecosystems theoretically and practically.