Identification and expression analysis of citrate synthase 3 gene family members in apple / 生物工程学报

As one of the key enzymes in cell metabolism, the activity of citrate synthase 3 (CS3) regulates the substance and energy metabolism of organisms. The protein members of CS3 family were identified from the whole genome of apple, and bioinformatics analysis was performed and expression patterns were analyzed to provide a theoretical basis for studying the potential function of CS3 gene in apple. BLASTp was used to identify members of the apple CS3 family based on the GDR database, and the basic information of CS3 protein sequence, subcellular localization, domain composition, phylogenetic relationship and chromosome localization were analyzed by Pfam, SMART, MEGA5.0, clustalx.exe, ExPASy Proteomics Server, MEGAX, SOPMA, MEME, WoLF PSORT and other software. The tissue expression and inducible expression characteristics of 6 CS3 genes in apple were determined by acid content and real-time fluorescence quantitative polymerase chain reaction (qRT-PCR). Apple CS3 gene family contains 6 members, and these CS3 proteins contain 473-608 amino acid residues, with isoelectric point distribution between 7.21 and 8.82. Subcellular localization results showed that CS3 protein was located in mitochondria and chloroplasts, respectively. Phylogenetic analysis divided them into 3 categories, and the number of genes in each subfamily was 2. Chromosome localization analysis showed that CS3 gene was distributed on different chromosomes of apple. The secondary structure of protein is mainly α-helix, followed by random curling, and the proportion of β-angle is the smallest. The 6 members were all expressed in different apple tissues. The overall expression trend from high to low was the highest relative expression content of MdCS3.4, followed by MdCS3.6, and the relative expression level of other members was in the order of MdCS3.3 > MdCS3.2 > MdCS3.1 > MdCS3.5. qRT-PCR results showed that MdCS3.1 and MdCS3.3 genes had the highest relative expression in the pulp of 'Chengji No. 1' with low acid content, and MdCS3.2 and MdCS3.3 genes in the pulp of 'Asda' with higher acid content had the highest relative expression. Therefore, in this study, the relative expression of CS3 gene in apple cultivars with different acid content in different apple varieties was detected, and its role in apple fruit acid synthesis was analyzed. The experimental results showed that the relative expression of CS3 gene in different apple varieties was different, which provided a reference for the subsequent study of the quality formation mechanism of apple.

Effect of Clomiphene Citrate on the Ultrastructure of Testis in Albino Rats

SUMMARY: The aim of the present work was to study the closer effect of clomiphene citrate on the ultrastructure of the testis of adult albino rats to provide a basis for optimizing this drug in the treatment of male infertility. The testes were removed from both groups under anesthesia and then prepared for examination by light using hematoxylin and eosin stains and a transmission electron microscope. Semithin sections were cut into 1 µm thick sections, stained with toluidine blue, and examined by light microscopy for a survey. The desired areas were placed in the center, and other areas were trimmed. Primary spermatocytes showed marked nuclear changes (pyknosis), and their nuclear membranes were ill-defined and disrupted. The cytoplasm showed widespread degeneration of mitochondria and lysosomes and focal degeneration of the rough endoplasmic reticulum compared with the control group. The spermatids were pale, and the two phases of spermatogenesis were distinctly identifiable in the control group but were confused in the treated group. Some spermatids had interrupted nuclear membranes, also containing degenerated mitochondria, focal fragmentation of rough endoplasmic reticulum, and free ribosomes. Spermatozoa in the treated group appeared deformed compared to the control, where they had deformed head caps. Leydig cells of the treated group have an irregularly shaped nucleus, with focal chromatin aggregation and peripheral chromatin condensation on the inner surface of the nuclear membrane. The observations of the present work indicate a possible causal relationship between testicular affection and ingestion of clomiphene citrate, which can be avoided by close medical observations using ultrasonography, semen analysis, or testicular biopsy to detect early malignant changes. Furthermore, the drug should not be used for more than three to six cycles and should be stopped for at least three cycles before reuse. When clomiphene citrate is ineffective in the treatment of male infertility, human menopausal gonadotropin (hMG) administration is typically selected. However, high-dose hMG therapy is associated with a variety of adverse effects. In this work, we report the success of a modified clomiphene citrate regimen in increasing sperm count without any hazards to the testicular tissue.

El objetivo del trabajo fue estudiar el efecto del citrato de clomifeno sobre la estructura de los testículos de la rata albina adulta, con la finalidad de determinar la mejor manera de utilizar este fármaco en el tratamiento de la infertilidad masculina. Los testículos se extrajeron bajo anestesia y para su análisis a través de microscopio de luz se tiñeron con HE. Además, las muestras fueron preparadas para su examen con microscopía electrónica de transmisión. Por otra parte, se cortaron secciones semifinas de 1 µm de espesor, se tiñeron con azul de toluidina y se examinaron mediante microscopía óptica. Los espermatocitos primarios mostraron cambios nucleares marcados (picnosis) y sus membranas nucleares estaban mal definidas y alteradas. En el grupo experimental las células presentaban el citoplasma con degeneración generalizada de las mitocondrias y de los lisosomas y una degeneración focal del retículo endoplásmico rugoso en comparación con el grupo control. Las espermátidas estaban pálidas y las dos fases de la espermatogénesis eran claramente identificables en el grupo control, pero se confundían en el grupo tratado. Algunas espermátidas tenían membranas nucleares interrumpidas, y también contenían mitocondrias degeneradas, fragmentación focal del retículo endoplásmico rugoso y ribosomas libres. Los espermatozoides del grupo tratado se presentaban deformados en comparación con el control. Las células de Leydig del grupo tratado presentaban un núcleo de forma irregular, con agregación focal de cromatina y condensación de cromatina periférica en la superficie interna de la membrana nuclear. Las observaciones del presente trabajo indican una posible relación causal entre la afección testicular y la ingestión de citrato de clomifeno, que puede evitarse mediante observaciones médicas minuciosas a través de ecografía, análisis de semen o biopsia testicular para detectar cambios malignos tempranos. Además, el medicamento no debiera ser usado durante más de tres a seis ciclos y debe suspenderse durante al menos tres ciclos antes de volver a usarlo. Cuando el citrato de clomifeno es ineficaz en el tratamiento de la infertilidad masculina, normalmente se selecciona la administración de gonadotropina menopáusica humana (hMG). Sin embargo, la terapia con hMG en dosis altas se asocia con una variedad de efectos adversos. En este trabajo, informamos el éxito de un régimen modificado con citrato de clomifeno para aumentar el recuento de espermatozoides sin riesgo para el tejido testicular.

Formulation and Evaluation of Orodispersible Tablet Containing Combination of Phosphodiesterase-5 Inhibitor and Herbal Aphrodisiac

Purpose: Sildenafil citrate is widely used drug for the treatment of Erectile Dysfunction (ED) and Ginseng is a natural aphrodisiac reported to benefit this condition. The objective of the present study was to develop orodispersible tablets (ODTs) containing combination of Sildenafil citrate and Ginseng extract to improve the bioavailability, reduce the dosing frequency and thereby maintaining the therapeutic efficacy of the drug. Methods: The ODTs were prepared using superdisintegrants such as Croscarmellose sodium (CCS), povidone, and sodium starch glycolate (SSG) at varying concentrations (2%, 4% and 6%) by direct compression. The bitter taste of Sildenafil citrate was masked by Doshion resin. The optimized formulation based on least disintegration time (DT) was chosen to reformulate using sublimating agents such as camphor, menthol or thymol at varying concentrations (1%, 2%, 3%) to further reduce the DT. The compatibility of drug with excipients was investigated and the prepared formulations were evaluated for pre and post-compression parameters. Results: The post-compression parameters such as weight variation, hardness, friability, DT and in-vitro drug release was found within specified limit. The formulation with camphor (2%) had DT of 12 sec and drug release >90% within 5 min hence was considered as optimized formulation. The accelerated stability study and kinetics modelling was performed for optimized formulation. Conclusion: The formulated Sildenafil citrate and Ginseng ODT’s were found to be promising formulation with quicker DT and drug release which will eventually have higher bioavailability and better efficacy along with averting the issues of swallowing and improving patient compliance.

Evaluation of the Effects of Aqueous Extract of Sarcocephalus latifolius on Sexual Behaviour in Male Rats

Background: Sarcocephalus latifolius, a plant of the Rubiaceae family, is found exclusively in tropical Africa, more particularly in sub-Saharan West Africa, stretching from Senegal to Zaire, but is also found as far east as Sudan (Sudan-Guinean zone). The leaves of this plant are used in traditional African medicine to treat numerous pathologies such as infertility. Aims/Objective: The present study aims to evaluate the effects of the aqueous extract of Sarcocephalus latifolius leaves on sexual parameters in male rats. Methods: The sexual parameters (latency of the mating position, latency of intromission, latency of ejaculation and mean copulation interval) were assessed by the classical method by administering distilled water, sildenafil citrate (reference molecule) called Viagra at the dose of 5 mg/kg of body weight and the aqueous extract of Sarcocephalus latifolius leaves at the dose of 1000 mg/kg of body weight per day for eight days to three batches of male rats respectively. These parameters were determined over a period of thirty minutes. Results: The results obtained show that the aqueous extract of Sarcocephalus latifolius leaves at a dose of 1000 mg/kg of body weight significantly (P < .001) decreases the latency time of mating position and the latency time of intromission, whereas this extract significantly (P < .001) increases the latency time of ejaculation and the average copulation interval. Conclusion: Aqueous extract of Sarcocephalus latifolius leaves has aphrodisiac properties which are due to sterols, flavonoids and saponosides.

A comparative study of the effects of citrate and heparin anticoagulation on coagulation function and efficacy in children with septic shock undergoing continuous blood purification / 中华危重病急救医学

Objective:To compare the effects of citrate and heparin anticoagulation on coagulation function and efficacy in children with septic shock undergoing continuous blood purification (CBP), and to provide guidance for CBP anticoagulation in children with septic shock.Methods:A case control study was conducted. Thirty-seven children with septic shock admitted to the pediatric intensive care unit (PICU) of the First Affiliated Hospital of Gannan Medical University from July 2019 to September 2022 were enrolled as the research subjects. The patients were divided into citrate local anticoagulation group and heparin systemic anticoagulation group according to different anticoagulation methods. The baseline data, the level of coagulation indicators [prothrombin time (PT), activated partial thromboplastin time (APTT), thrombin time (TT), fibrinogen (Fib), D-dimer] before treatment and 1 day after weaning from CBP, serum inflammatory mediators [interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), hypersensitivity C-reactive protein (hs-CRP), procalcitonin (PCT)], bleeding complications during CBP and 7-day mortality were collected.Results:A total of 37 cases were enrolled finally, including 17 cases with citric acid local anticoagulation and 20 cases with heparin systemic anticoagulation. There was no statistically significant difference in general data such as gender, age, and body weight of children between the two groups. There were no statistically significant differences in baseline levels of coagulation indicators and inflammatory mediators before treatment of children between the two groups. One day after weaning from CBP, both groups showed varying degrees of improvement in coagulation indicators compared with those before treatment. Compared with before treatment, the PT of the heparin systemic anticoagulation group was significantly shortened after 1 day of weaning (s: 11.82±2.05 vs. 13.64±2.54), APTT and TT were significantly prolonged [APTT (s): 51.54±12.69 vs. 35.53±10.79, TT (s): 21.95±4.74 vs. 19.30±3.33], D-dimer level was significantly reduced (mg/L: 1.92±1.58 vs. 4.94±3.94), with statistically significant differences (all P < 0.05). While in the citrate local anticoagulation group, only APTT was significantly prolonged after treatment compared with that before treatment (s: 49.28±10.32 vs. 34.34±10.32, P < 0.05). There were no statistically significant differences in other coagulation indicators compared with before treatment. Compared with the citric acid local anticoagulation group, the PT of the heparin systemic anticoagulation group was significantly shortened after treatment (s: 11.82±2.05 vs. 13.61±3.05, P < 0.05), and the D-dimer level was significantly reduced (mg/L: 1.92±1.58 vs. 3.77±2.38, P < 0.01). The levels of inflammatory mediators in both groups were significantly reduced 1 day after CBP weaning compared with those before treatment [citric acid local anticoagulation group: hs-CRP (mg/L) was 12.53±5.44 vs. 22.65±7.27, PCT (μg/L) was 1.86±1.20 vs. 3.30±2.34, IL-6 (ng/L) was 148.48±34.83 vs. 202.32±48.62, TNF-α (ng/L) was 21.38±7.71 vs. 55.14±15.07; heparin systemic anticoagulation group: hs-CRP (mg/L) was 11.82±4.93 vs. 21.62±8.35, PCT (μg/L) was 1.90±1.08 vs. 3.18±1.97, IL-6 (ng/L) was 143.81±33.41 vs. 194.02±46.89, TNF-α (ng/L) was 22.44±8.17 vs. 56.17±16.92, all P < 0.05]. However, there was no statistically significant difference between the two groups (all P > 0.05). There was no statistically significant difference in bleeding complication during CBP and 7-day mortality in children between the citrate local anticoagulation group and the heparin systemic anticoagulation group (5.9% vs. 30.0%, 17.6% vs. 20.0%, both P > 0.05). Conclusions:Heparin for systemic anticoagulation and regional citrate anticoagulation can significantly reduce the levels of IL-6, TNF-α, hs-CRP and PCT in children with septic shock, and relieve inflammatory storm. Compared with citric acid local anticoagulation, heparin systemic anticoagulation can shorten the PT and reduce the level of D-dimer in children with septic shock, which may benefit in the prevention and treatment of disseminated intravascular coagulation (DIC).

Expression and Clinical Significance of ATP Citrate Lyase in Hepatocellular Carcinoma / 中国医学科学院学报

Objective To investigate the role of ATP citrate lyase(ACLY)in the development of hepatocellular carcinoma(HCC)and the impact of this enzyme on the immune microenvironment of HCC.Methods We utilized the University of Alabama at Birmingham Cancer Data Analysis Portal and the Gene Expression Profiling Interactive Analysis to identify the changes in ACLY expression and prognosis across different tumor types from The Cancer Genome Atlas.With HCC as the disease model,we analyzed the ACLY expression in HCC samples from the gene expression database.Furthermore,we collected the clinical specimens from HCC patients to verify the mRNA and protein levels of ACLY.In addition,we conducted transcriptome sequencing after knocking down the expression of ACLY to analyze the differentially expressed genes and investigated the impact of ACLY expression interference on cell proliferation and other functions.Finally,we explored the correlations of ACLY with immune cells and immune infiltration in the tumor microenvironment,new antigens,and immune checkpoint genes.Results ACLY expression was significantly up-regulated in solid tumors including HCC(all P<0.05),and high ACLY expression was associated with overall survival rate in HCC(P=0.005).Furthermore,high ACLY expression affected the presence of immune cells(e.g.,tumor-associated fibroblasts)and the expression of genes involved in lipid metabolism(all P<0.05).Conclusions ACLY is closely related to the occurrence and development of HCC and lipid metabolism abnormalities.Moreover,it has a specific impact on the immune microenvironment of HCC.

Development of the novel ACLY inhibitor 326E as a promising treatment for hypercholesterolemia

Hepatic cholesterol accumulation is an important contributor to hypercholesterolemia, which results in atherosclerosis and cardiovascular disease (CVD). ATP-citrate lyase (ACLY) is a key lipogenic enzyme that converts cytosolic citrate derived from tricarboxylic acid cycle (TCA cycle) to acetyl-CoA in the cytoplasm. Therefore, ACLY represents a link between mitochondria oxidative phosphorylation and cytosolic de novo lipogenesis. In this study, we developed the small molecule 326E with an enedioic acid structural moiety as a novel ACLY inhibitor, and its CoA-conjugated form 326E-CoA inhibited ACLY activity with an IC50 = 5.31 ± 1.2 μmol/L in vitro. 326E treatment reduced de novo lipogenesis, and increased cholesterol efflux in vitro and in vivo. 326E was rapidly absorbed after oral administration, exhibited a higher blood exposure than that of the approved ACLY inhibitor bempedoic acid (BA) used for hypercholesterolemia. Chronic 326E treatment in hamsters and rhesus monkeys resulted in remarkable improvement of hyperlipidemia. Once daily oral administration of 326E for 24 weeks prevented the occurrence of atherosclerosis in ApoE-/- mice to a greater extent than that of BA treatment. Taken together, our data suggest that inhibition of ACLY by 326E represents a promising strategy for the treatment of hypercholesterolemia.

Pathologically Elevated-Cyclic Stretch Suppressed Vascular Smooth Muscle Cell Mitochondrial Biogenesis by Down-regulating PGC1α Expression / 医用生物力学

Objective To investigate the effect of pathologically elevated-cyclic stretch induced by hypertension on mitochondrial biogenesis of vascular smooth muscle cells (VSMCs), and the role of PGC1α in this process. Methods The Flexcell-5000T stretch loading system in vitro was applied to VSMCs with a frequency of 1. 25 Hz and an amplitude of 5% or 15% to simulate the mechanical environment under normal physiological or hypertensive pathological conditions respectively. Western blotting and qPCR were used to detect the expression of PGC1α, citrate synthase and mitochondrial DNA (mtDNA) copy number in VSMCs under normal physiological or hypertensive pathological conditions. VSMCs were treated with PGC1α specific activator ZLN005 to promote PGC1α expression or specific interfering fragment siRNA to inhibit PGC1α expression in order to detect the effect on citrate synthase and mtDNA copy number. Results Compared with 5% physiological cyclic stretch, 15% pathologically elevated-cyclic stretch significantly suppressed the expression of PGC1α, citrate synthase and mtDNA copy number in VSMCs. Compared with control group, the protein expression of PGC1α was significantly decreased and increased respectively. When VSMCs transfected with PGC1α siRNA or incubated PGC1α activator ZLN005, the expression of citrate synthase and mtDNA copy number were also significantly down regulated and up-regulated in VSMCs accordingly. Under physiological cyclic stretch conditions, the protein level of PGC1α was significantly down-regulated by PGC1α siRNA, which also significantly down-regulated citrate synthase expression and mtDNA copy number. The protein expression of PGC1α was significantly up-regulated by ZLN005, which also enhanced the expression of citrate synthase and mtDNA copy number. Conclusions The pathological cyclic stretch induced by hypertension significantly down-regulated the expression of citrate synthase and mtDNA copy number via suppressing the expression of PGC1α, resulting in mitochondrial dysfunction of VSMCs. PGC1α may be a potential therapeutic target molecule to alleviate the progression of hypertension.

Citrate does not Change Viscoelastic Haemostatic Assays after Cardiopulmonary Bypass

Context:Viscoelastic hemostatic assays (VHA) are commonly used to identify specific cellular and humoral causes for bleeding in cardiac surgery patients. Cardiopulmonary bypass (CPB) alterations to coagulation are observable on VHA. Citrated VHA can approximate fresh whole blood VHA when kaolin is used as the activator in healthy volunteers. Some have suggested that noncitrated blood is more optimal than citrated blood for point?of?care analysis in some populations. Aims: To determine if storage of blood samples in citrate after CPB alters kaolin activated VHA results. Settings and Design: This was a prospective observational cohort study at a single tertiary care teaching hospital. Methods and Material: Blood samples were subjected to VHA immediately after collection and compared to samples drawn at the same time and stored in citrate for 30, 90, and 150 min prior to kaolin activated VHA both before and after CPB. Statistical Analysis Used: VHA results were compared using paired T?tests and Bland–Altman analysis. Results: Maximum clot strength and time to clot initiation were not considerably different before or after CPB using paired T?tests or Bland– Altman Analysis. Conclusions: Citrated samples appear to be a clinically reliable substitute for fresh samples for maximum clot strength and time to VHA clot initiation after CPB. Concerns about the role of citrate in altering the validity of the VHA samples in the cardiac surgery population seem unfounded.

Diagnostic Value of D-Dimer and INR in Patients Suspected to Have Prosthetic Valve Dysfunction

ABSTRACT Introduction: Prosthetic valve dysfunction is a potentially critical complication of heart valve replacement. An easy and quickly applicable diagnostic procedure is required for recognizing the prosthetic valve dysfunction. The purpose of this study was to prospectively define the diagnostic value of D-dimer and INR level in predicting prosthetic valve dysfunction. Methods: This cross-sectional study was performed in 70 patients suspected to have prosthetic valve dysfunction admitted to Imam Ali Hospital, affiliated with Kermanshah University of Medical Sciences (KUMS), Kermanshah Province, Iran. Cinefluoroscopy, as the gold standard diagnostic test, was used for the diagnosis of prosthetic valve dysfunction in enrolled patients. Two milliliters of blood from each patient were taken into a tube containing sodium citrate anticoagulant. To evaluate D-dimer, the cutoff value was set at 500 ng/ml. Also, to evaluate international normalized ratio (INR), the cutoff value was set at 2. Sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), positive likelihood ratio (PLR), and negative likelihood ratio (NLR) of the serum markers were used to describe predictive properties. Results: Of 70 patients, 27 (38.6%) were male and 43 (61.4%) were female, and the mean age was 54.67±15.11 years (range, 18 to 80 years). Of 70 patients, 27 (38.6%) had prosthetic heart valve malfunction demonstrable by fluoroscopy, and 19 patients (27.1%) had D-dimer levels >500 ng/ml. Elevated D-dimer levels (>500 ng/ml) have been indicated to have sensitivity of 70.4%, and hence an NPV of 84.3%, specificity of 100%, PPV of 100%, NLR of 0.3, and the infinity value of PLR for predicting prosthetic valve dysfunction. There was a significant relationship between fluoroscopy and D-dimer test (P=0.001). A kappa coefficient value of 0.745 indicated a substantial agreement between D-dimer and fluoroscopy testing. Mixing test (combination of D-dimer and INR) showed to have 100% sensitivity, and hence a NPV of 69.8%, specificity of 69.8%, PPV of 51.8%, NLR of 1.41, and PLR of 1.44 for predicting prosthetic valve dysfunction. Conclusion: D-dimer with moderate sensitivity and high specificity is an ideal marker for the diagnosis of prosthetic valve dysfunction in suspected patients. Enhanced plasma D-dimer level is not by itself diagnostic of a prosthetic valve dysfunction but may alert physicians to refer the patient for more detailed examination, preferably by fluoroscopy. Mixing test with 100% sensitivity can apply as a rule-out test.

Efectividad de anticoagulación regional con citrato en terapia de reemplazo renal continua / Effectiveness of regional citrate anticoagulation in continuous renal replacement therapy

BACKGROUND: Anticoagulation in continuous renal replacement therapy (CRRT) is essential to counteract the coagulation cascade activation, induced by the dialysis circuit. Heparin is the most widely used anticoagulant, followed by regional citrate anticoagulation (RCA). AIM: To determine the effectiveness and safety of anticoagulant treatment with citrate in CRRT. Material and Methods: Retrospective study of adults in CRRT hospitalized between the years 2014 and 2020 in critical units, who required change to RCA according to established protocols. RESULTS: We studied 24 patients aged 63 ± 13 years (12 females). The reasons for admission were acute kidney injury (AKI) in 80% and stage 5 chronic kidney disease in 20%. The indication of RCA in 75% of patients was by coagulation of more than 3 circuits in 24 hours. The duration of the circuit in RCA was 18.5 ± 4.8 hours versus 11.9 ± 4.9 hours with heparin (p < 0.0001). There were 19 mild complications that did not affect the RCA. Conclusions: RCA is feasible to perform, it is a safe and efficient procedure if it is protocolized, allowing a longer duration of the dialysis circuit.

Treatment of Infertility in Women with Polycystic Ovary Syndrome in a Single Center Study

Background: Anovulatory infertility is caused by polycystic ovarian syndrome in 80 percent of patients. Preconception guidelines, such as lifestyle modification (weight loss) to avoid fetal neural tube abnormalities, and quitting smoking and drinking alcohol, are all part of the early treatment. A clomiphene citrate medication for timed intercourse is the first-line pharmacological treatment for producing ovulation. Exogenous gonadotropins or laparoscopic ovarian surgery are two options for second-line pharmaceutical treatment (ovarian drilling). Ovulation induction using clomiphene citrate or gonadotropins is effective, with a 70 % cumulative live birth rate. When laparoscopy is necessary, ovarian drilling should be done; this operation is usually successful in around half of the instances. Finally, when the previous interventions have failed, a high-complexity reproduction treatment (in vitro fertilization or intracytoplasmic sperm injection) is recommended. There is no evidence that metformin should be used routinely in the treatment of infertility in anovulatory women with polycystic ovary syndrome. Aromatase inhibitors show promise, but more research is needed to confirm their safety.Methods:This study was conducted in Department of gynecology and obstetrics, Dhaka Medical College Hospital, Dhaka, from January 2019 to December 2019. A total number of 100 patients with multiple myeloma were analyzed cytogenetically by interphase fluorescence in situ hybridization (iFISH). The collected data were analyzed by using the Statistical Package for Social Science (SPSS-24) for windows version 24.0.Conclusion:PCOS is a frequent syndrome and the most frequent cause of infertility. PCOS is defined as a syndrome with at least two of three of the Rotterdam criteria. A complete evaluation of the infertility is needed to exclude other causes of infertility..

Optimized synthesis process of tofacitinib citrate / 中国药科大学学报

@#In order to improve the yield and simplify the operation, the synthesizing process of JAK3 inhibitor tofacitinib citrate was improved based on the analysis of the methods previously published.Using 2, 4-dichloro-7H-pyrrolo [2, 3-d] pyrimidine and (3R, 4R)-1-benzyl-N, 4-dimethylpiperidin-3-amine dihydrochloride as starting materials, tofacitinib citrate was obtained through four steps of nucleophilic substitution, catalytic transfer hydrogenation, cyanide acetylation and citrate salt, and its crystal form was consistent with the original research.After optimization, the yield was better than those reported in literature, and the mild reaction conditions were suitable for industrial production.

Research progress on caffeine citrate in the treatment of neonatal respiratory distress syndrome / 中国小儿急救医学

Neonatal respiratory distress syndrome is typically characterized by progressive exacerbation of respiratory distress shortly after birth, which is more common in preterm infants and has a high disability and mortality rate.Caffeine citrate has been used in the treatment of premature infants with respiratory distress syndrome to enhance the contraction of the diaphragm and optimize the function of respiratory muscles to accelerate the recovery of spontaneous breathing.This review summarized the use of caffeine citrate in premature infants with respiratory distress syndrome.

Research progress on caffeine citrate in the treatment of neonatal respiratory distress syndrome / 中国小儿急救医学

Neonatal respiratory distress syndrome is typically characterized by progressive exacerbation of respiratory distress shortly after birth, which is more common in preterm infants and has a high disability and mortality rate.Caffeine citrate has been used in the treatment of premature infants with respiratory distress syndrome to enhance the contraction of the diaphragm and optimize the function of respiratory muscles to accelerate the recovery of spontaneous breathing.This review summarized the use of caffeine citrate in premature infants with respiratory distress syndrome.

Metabolic dysregulation and emerging therapeutical targets for hepatocellular carcinoma

Hepatocellular carcinoma (HCC) is an aggressive human cancer with increasing incidence worldwide. Multiple efforts have been made to explore pharmaceutical therapies to treat HCC, such as targeted tyrosine kinase inhibitors, immune based therapies and combination of chemotherapy. However, limitations exist in current strategies including chemoresistance for instance. Tumor initiation and progression is driven by reprogramming of metabolism, in particular during HCC development. Recently, metabolic associated fatty liver disease (MAFLD), a reappraisal of new nomenclature for non-alcoholic fatty liver disease (NAFLD), indicates growing appreciation of metabolism in the pathogenesis of liver disease, including HCC, thereby suggesting new strategies by targeting abnormal metabolism for HCC treatment. In this review, we introduce directions by highlighting the metabolic targets in glucose, fatty acid, amino acid and glutamine metabolism, which are suitable for HCC pharmaceutical intervention. We also summarize and discuss current pharmaceutical agents and studies targeting deregulated metabolism during HCC treatment. Furthermore, opportunities and challenges in the discovery and development of HCC therapy targeting metabolism are discussed.

Magnesium citrate inhibits hyperphosphorus-induced VSMCs calcification through activating calcium-sensitive receptor / 西安交通大学学报(医学版)

【Objective】 To investigate the inhibitory effect of magnesium citrate （MgCit） on hyperphosphorus-induced VSMCs calcification and its mechanisms. 【Methods】 VSMCs were divided into the following groups： normal control group， high-phosphorus group， low-dose MgCit group， high-dose MgCit group and high-dose MgCit+NPS2143 （calcium-sensitive receptor inhibitor） group. Alizarin red staining was used for semi-quantitative analysis of VSMCs calcification and the calcium content in VSMCs was detected by the commercial kit. The osteogenic transdifferentiation parameters including alkaline phosphatase （ALP） activity， as well as mRNA and protein levels of smooth muscle 22α （SM22α）， runt-related transcription factor 2 （RUNX2） and bone morphogenetic protein 2 （BMP2）， were detected in each group. 【Results】 Compared with those in the control group， the calcium content of VSMCs in the model group was increased， ALP activity， and the mRNA and protein expressions of RUNX2 and BMP2 were increased， and the mRNA expression of SM22α was decreased （P<0.05）. MgCit could reduce VSMCs calcification， decrease ALP activity， decrease mRNA and protein levels of RUNX2 and BMP2， and increase mRNA levels of SM22α induced by high-phosphorous （P<0.05）. The above effects of MgCit were reduced when NPS2143 was administered simultaneously （P<0.05）. 【Conclusion】 MgCit can reduce VSMCs calcification and osteogenic transdifferentiation induced by high phosphorus through activating of CaSR.

How some biochemical indexes of apheresis platelet donors were affected by different calcium supplement methods at blood donation intervals / 中国输血杂志

【Objective】 To investigate the effect of two different oral calcium supplementation methods(method 1: bone health basic supplements in the interval plus routine calcium supplement; method 2: only routine calcium supplement on the day of donation)on platelet donors, to improve some biochemical indexes of the body caused by citrate anticoagulant. 【Methods】 A total of 252 long-term regular platelet donors in our station from January 2019 to December 2020 were selected and randomly divided into two groups: experimental group (n=126) were given 10% calcium gluconate oral solution 20 mL before platelet collection, orally. Oral caltrate D3 (containing 600 mg calcium carbonate and 125 international units of vitamin D3) was administered in the interval (20±2 days), 1 tablet per day. Oral alfacalcidol soft capsule (containing alfacalcidol 0.25 μg), 1 tablet per day; control group (n=126) were given routine administration of 10% calcium gluconate oral solution 20 mL before platelet collection. Serum PTH and serum concentrations of calcium, magnesium, phosphorus and 25 hydroxyvitamin D of the two groups were detected at 0 min after injection and 0 min after the end of the last circulation, respectively. Meanwhile, the occurrence of blood donation reactions was observed in the two groups. 【Results】 In the experimental group and control group, serum PTH concentration both increased significantly after donation, while serum calcium, inorganic phosphorus, magnesium and 25 hydroxyvitamin D decreased significantly(P<0.01). After donation, 4 donors in the experimental group and 8 in the control group showed citrate reaction, with the incidence of adverse reactions at 3.17%(4/126)and 6.34%(8/126), respectively. Before blood collection, serum calcium concentration in the experimental group was higher than that in the control group, while PTH concentration was lower than that in the control group (P<0.01), but no significant differences were noticed in serum magnesium, inorganic phosphorus and 25 hydroxyl vitamin D concentrations (P>0.05), and so was in the control group after blood collection. 【Conclusion】 Method 1 can further slow down the increase of serum PTH and the decrease of serum calcium, and can effectively reduce the citrate response of blood donors compared with method 2, which is of great significance to prevent osteoporosis in long-term regular blood donors.

Effects of low molecular weight heparin sodium and sodium citrate on hemodialysis patients at high risk of bleeding: a comparative study / 中国输血杂志

【Objective】 To compare the therapeutic effects of low molecular weight heparin sodium and sodium citrate on hemodialysis(HD) patients at high risk of bleeding. 【Method】 A total of 96 patients at high risk of bleeding on maintenance hemodialysis from May 2018 to May 2020 were enrolled and divided randomly into control group(n=48) and observation group(n=48). Patients in control group received systemic anticoagulation with heparin, and patients in observation group adopted regional citrate anticoagulation of in vitro dialyzer. The indexes of prothrombin time (PT), activated partial thromboplastin time (APTT), fibrinogen (Fg), D-dimer, service time of filters and the dialysis efficiency, as well as the complication occurrences of clotting events, bleeding, metabolic alkalosis and hypocalcaemia were compared between the two groups. 【Results】 No significant difference was found in coagulation indicators (ACT, PT, Fg, APPT and D-dimer) before and after hemodialysis in observation group(P>0.05), but those in control group were prolonged significantly (P0.05). The incidence of filter clotting (0.0%), dialyzer pipeline clotting (0.0%) and bleeding (2.1%) in observation group were less than those in control group (8.3%, 6.2%, 14.6%, respectively) (P0.05). 【Conclusion】 Regional anticoagulation with citric acid in vitro is safe for patients on maintenance hemodialysis at high risk of bleeding, and can greatly prolong service time of filters, reduce bleeding and thrombocytopenia rates, bring less effects to coagulation function in vivo.

Efeitos do citrato de clomifeno, letrozol e estradiol sobre apoptose em células do cumulus oophorus humano / Effects of clomiphene citrate, letrozole and estradiol on apoptosis in human cumulus oophorus cells

Citrato de clomifeno (CC) e letrozol (LE) são indutores de ovulação que, apesar das altas taxas de ovulação confirmada, atingem baixas taxas de gravidez. Este estudo teve como objetivo investigar os efeitos de CC e LE in vitro, isoladamente ou em combinação com estradiol (E), na apoptose de células do cumulus oophorus humano. Realizamos um estudo prospectivo controlado utilizando culturas primárias de células do cumulus de pacientes submetidas à fertilização in vitro (n=22). A coloração com Giemsa e a imunocitoquímica para alfa-inibina foram utilizadas para avaliar a pureza e a morfologia da cultura celular. A viabilidade celular foi avaliada pelo ensaio MTT, o ciclo celular por citometria de fluxo e a expressão gênica de Caspase-3, Bax e Superóxido dismutase 2 (SOD-2) e S26 por reação em cadeia de polimerase (PCR) em tempo real. As células foram tratadas por 24 horas em 5 grupos de tratamento: CC, CC + E, LE, LE + E e controle. Nenhum dos tratamentos afetou a viabilidade celular, mas o LE reduziu a porcentagem média de células na fase S em relação ao controle (24,79 versus 21,70, p=0,0014). O tratamento com CC aumentou a expressão gênica de Bax (4 vezes) e SOD-2 (2 vezes), que foi revertida quando adicionado E à cultura. A expressão de SOD-2 aumentou em células tratadas com LE quando comparado ao controle (4 vezes), que foi também revertida por adição de E. Estes achados sugerem que CC e LE não afetam significativamente a viabilidade das células do cumulus humana. Porém, houve modulação na expressão de genes envolvidos na apoptose por essas drogas isoladamente e em associação com E, sugerindo que CC e LE podem ter efeitos diretos nas células do cumulus além de seus mecanismos de ação conhecidos.

Clomiphene citrate (CC) and letrozole (LE) are ovulatory stimulants that, despite high ovulation rates, achieve low pregnancy rates. This study aimed to investigate the in vitro effects of CC and LE, alone or in combination with estradiol (E), on apoptosis in human cumulus cells. We performed a controlled prospective study using primary cumulus cell cultures from patients undergoing in vitro fertilization (n=22). Giemsa stain and alpha-inhibin immunocytochemistry was used to assess cell culture purity and morphology. Cell viability was evaluated by MTT assay, cell cycle status by flow cytometry, and Caspase-3, Bax and superoxide dismutase 2 (SOD-2), and S26 gene expression by real-time polymerase chain reaction (qPCR). Cells were treated for 24 hours in 5 conditioned media: CC, CC + E, LE, LE + E and control. None of the treatments affected cell viability, but LE reduced the mean percentage of cells in the S phase compared to control (24.79 versus 21.70, p=0.0014). CC treatment increased mRNA expression of Bax (4 fold) and SOD-2 (2 fold), which was reversed by co-treatment with E. SOD-2 expression increased in cells treated with LE compared to control (4 fold), which was also reversed by E. These findings suggest that CC and LE do not significantly affect the viability of human cumulus cells. Still, the expression of genes involved in apoptosis was modulated by these drugs alone and in association with E, suggesting that CC and LE may have direct effects on cumulus cells beyond their known mechanisms of action.