[The research of sentinel lymph node biopsy after neoadjuvant chemotherapy for breast cancer patients].

With the extensive use of sentinel lymph node biopsy (SLNB), some breast cancer patients could avoid axillary lymph node dissection (ALND) and its complications. Neoadjuvant chemotherapy plays an important role in the multimodality therapies of breast cancer. After neoadjuvant chemotherapy, some patients with breast cancer were down-staged from positive axillary lymph node (cN+ ) to clinically negative (cN0). For these patients, the feasibility and safety of sentinel lymph node biopsy remains controversial. However, with the application of new technologies, SLNB is expected to become the main treatment for breast cancer patients with stage cN0 after neoadjuvant chemotherapy.

[In vitro fertilization-embryo transfer affects focal adhension kinase signaling pathway in early placenta].

OBJECTIVE: To study the effects of in vitro fertilization-embryo transfer (IVF-ET) technique on gene expression of focal adhension kinase (FAK) signaling pathway in early placental trophoblast cells, and to explore the effects of IVF-ET technology on the development and function of early placenta. METHODS: We collected 7-8 weeks of gestation placenta tissue as a study group by ultrasound guided reduction of fetal from double embryo transfer under IVF-ET technology. In the control group, placenta tissues were obtained from the spontaneous abortion of natural pregnancy twin 7-8 weeks. Microarray hybridization analysis was performed on the placenta tissue of the two groups using the Affymetrix HG-U133 Plus 2.0 gene chip. Eight differentially expressed genes were identified by real-time quantitative polymerase chain reaction (qRT-PCR), and unsupervised clustering analysis and functional bioinformatics analysis were performed for the differentially expressed genes. RESULTS: Twenty-eight cases of IVF-ET reduced fetal villi and 8 cases of spontaneous abortion villi were collected. A total of 8 placental villi were detected by the gene chip. Compared with the natural pregnancy control group, 32 differentially expressed genes in the placental FAK signaling pathway were expressed in IVF-ET. The differential expression was greater than or equal to 2 times, of which 12 genes were up-regulated and 20 were down-regulated. The qRT-PCR showed that the expression of the 8 genes in FAK signaling pathways of IVF-ET was significantly different from that in the placenta of natural pregnancy, which was consistent with the result of the gene chip detection. The FAK signal pathway gene localization showed that the FAK gene was mainly located in the upstream of the signal pathway in the placenta of IVF-ET. The placental trophoblast cells maintained the FAK signaling pathway function through gene expression compensation. CONCLUSION: There are gene expression differences in the FAK signaling pathway between the IVF-ET derived early placenta and the natural pregnancy placenta. The differentially expressed genes are involved in many key functions of the FAK signaling pathway and affect the early development and function of the IVF-ET placenta, while the placental trophoblast cells change gene expression for interference to compensate for IVF-ET technology itself, maintain normal function of the FAK signaling pathway, and satisfy the need for placental and fetal development.

[Immunological damage effects of cytomegalovirus infection on bone marrow hematopoietic cells].

Objective: Toobserve the damage effects of cytomegalovirus (CMV) infection on hematopoietic cells and to investigate the clinical significance. Methods: (1) ELISA assay wasused to detect IL-17 and IFN-γ levels in the peripheral blood serum of 36 patients on pretherapy and posttherapy. (2) Changes of peripheral blood lymphocyte subsets were detected by FACS. (3) Cytological observation of cervical lymph nodes was executed by needle aspiration cytology. (4) Cellular immunochemistry and immunofluorescence staining were performed to observe the POX release, HLA-DR expression, IL-17A and IFN-γ secretion-like expression status of activated immune cells in the bone marrow hematopoietic microenvironment. Serum samples from healthy individuals were used as controls. Bone marrow smears from patients without iron deficiency anemia were compared as bone marrow immunostaining. Results: (1) Serum levels of IL-17 and IFN-γ were significantly increased in CMV-infected patients [IL-17 (48.23±3.86) ng/L vs (20.16±1.05) ng/L,respectively; IFN-γ (40.16±3.11) vs (8.17±1.92) ng/L,P<0.01]. (2) The proportion of CD16+/CD56+NK cells was significantly increased in patients [(43.54±6.01)% vs (14.01±3.25)%, P<0.01]. The proportion of CD3+CD4+T and CD3+CD8+T cells decreased,(20.91±53.15)% vs (35.10±4.88)%, and (15.41±5.13)% vs (25.11±3.92)%,respectively,P<0.05. (3) Large numbers of abnormal lymphocytes and macrophages (MΦ) that engulf large quantities of CMV inclusion bodies were observed in bone marrow and lymph nodes. CMV infected and destroied the hematopoietic cells of various lines in the bone marrow. The activated MΦ phagocytizedthe CMV inclusion bodies and also phagocytosed CMV-infected blood cells. (4) Activated MΦ in bone marrow hematopoietic microenvironment released POX strongly positive, highly expressed class Ⅱ HLA-DR, and highly expressed inflammatory factors IL-17A and IFN-γ. (5) Twenty-twopatients with elevated WBC were treated with ganciclovir combined with antibiotics for 2 weeks after the disappearance of the foci, WBC counts and CMV-IgM levels of 16 cases were reduced to normal.Six patients with CMV who were not turned negative were tprolonged,and the granulocyte and/or platelet counts fell below normal range. Fourteenpatients withreduced granulocyte or platelet count, CMV-IgM levels were slow descend,and the ganciclovir treated more than 4 weeks. Conclusions: CMV can infect hematopoietic bone marrow nucleated blood cells and destroy hematopoiesis. NK and MΦ cells are important effector cells against CMV infection. Activated macrophages are dual in nature, they can engulf CMV virus and virus-infected blood cells,and also aggravate bone marrow immune damage by releasing inflammatory factors such as POX and IL-17A and IFN-γ.

[Study on the spatial expression of trophectoderm cells in human embryonic prenatal blastocysts].

OBJECTIVE: To study the spatial expression of trophectoderm cells in human embryonic preantral blastocysts. METHODS: The study used Gardner score 5AA blastocysts harvested on day 6 after fertilization from assisted reproductive technology. Microcapsules were used to separate trophectoderm cells from the epidermal cells. Single-cell sequencing was performed. P<0.05 was calculated by unpaired t test, and the difference was 2 times. Here we determined, for the first time, global gene expression patterns in the polar/mural trophectoderm isolated from human blastocysts. Unsupervised hierarchical clustering analysis and gene ontology (GO) functional classification were performed using bioinformatics software. Differentially expressed genes were annotated by the Database for Annotation, Visualization and Integrated Discovery. Functions of differentially expressed genes were further annotated using encyclopedia of genes and genomes. RESULTS: The results showed that there were up to 306 genes in the trophoblast cells and up to 75 genes in the trophoblast cells. Unsupervised cluster analysis of polar trophoblast cells and mural trophoblast cells were divided into two groups, belonging to different types and biological functions. Differences in gene function indicated that the biological functions of GO gene uptake genes were mainly transcription, energy metabolism, protein synthesis, transport, oxidative stress, ion transport, protein synthesis and transport, cell cycle regulation, actin growth, etc. They were mainly involved in ubiquitin-mediated protein hydrolysis, oxidative phosphorylation, Wnt signaling pathway, estrogen androgen metabolism and other signal pathways; wall trophoblast cells up-regulated gene GO biological function, which was mainly proteolytic metabolism, cell cycle arrest, apoptosis, activation of MAPK, carbohydrate transport, synaptic regulation, cell growth, calcium channel activation, positive B cell differentiation, T cell apoptosis and other biological functions, which were mainly involved in B cell receptor, T cell receptor, white blood cells cross-endothelial transplantation, VEGF expression, gap connection, GnRH secretion, apoptosis and other signaling pathways. CONCLUSION: The gene expression of blastocysts trophectoderm is revealed from the spatial dimension, indicating that differentiation of polar and mural trophectoderm of blastocysts is accompanied by differences between the two cell lineages, and the polar and mural trophectoderms are coordinated with each other and the blastocyst hatching and embryo implantation processes are finely adjusted. Further data analysis is expected to find the endogenous molecular specificity of the regulation of embryo implantation.

Tanshinone IIA ameliorates lead (Pb)-induced cognitive deficits and oxidative stress in a rat pup model.

OBJECTIVE: Chronic lead (Pb) exposure affects the developing central nervous system, whereas Tanshinone IIA (TSA) improves cognitive deficits. METHODS: In this study, we investigated the effects of TSA against lead-induced neurotoxicity in a rat pup model. A total of thirty two healthy male Wistar rats were randomly divided into four groups: lead-treated group, lead plus TSA-treated 1 group, lead plus TSA-treated 2 group, and controls. After a 4-week lead exposure, memory function was determined using Morris water maze and the concentration of lead was measured in blood. Total superoxide dismutase (T-SOD), glutathione (GSH), malonaldehyde (MDA) and brain-derived neurotrophic factor (BDNF) activities were determined in hippocampus samples. RESULTS: Lead exposure causes decrease of body weight; increase of the blood lead concentration; decrease of antioxidant activities and BDNF content. However, co-administration of TSA with lead ameliorated the weight loss. Furthermore, TSA inhibited neurotoxicity as evidenced by decreased latency period and increase in percentage of time spent in the target quadrant. Administration of TSA also improved antioxidant activities by increased T-SOD, GSH, and decreased MDA activities compared to lead-treated group. CONCLUSION: This study provides evidence of that TSA has a neuroprotective effect against lead-induced cognitive deficit by enhancing antioxidant activities in the brain (Tab. 2, Fig. 3, Ref. 27).

Resistin expression in adipose tissues and its effect on glucose metabolism in rats with brain injury.

Resistin (RSTN) expression in subcutaneous adipose tissue, and its effect on glucose metabolism in rats with traumatic brain injury, was investigated using real-time PCR, western blots, and enzyme linked immunoassays. Our results show that the expression of RSTN mRNA (3.192 ± 0.046, 4.016 ± 0.010, 6.004 ± 0.020, 8.213 ± 0.013, 11.199 ± 0.174, 15.094 ± 0.030), protein levels (1.79 ± 0.05, 1.98 ± 0.07, 2.75 ± 0.08, 3.19 ± 0.08, 4.25 ± 0.11, 4.48 ± 0.07), levels of serum insulin (512.96 ± 1.21, 580.57 ± 1.52, 769.71 ± 2.22, 826.08 ± 2.03, 1262.25 ± 3.40, 1512.80 ± 3.93), and fasting blood glucose levels (10.277 ± 0.040, 12.776 ± 0.038, 13.403 ± 0.263, 14.698 ± 0.100, 16.637 ± 0.110, 19.416 ± 0.025) were significantly higher in the traumatic rat group compared to the control group (P < 0. 05). Quantitative insulin sensitivity check index (QUICKI) was significantly lower in the traumatic group (-8.570 ± 0.005, -8.912 ± 0.004, -9.241 ± 0.022, -9.404 ± 0.007, -9.952 ± 0.007, -10.288 ± 0.002) than in the control group (-7.633 ± 0.003, -7.639 ± 0.004, -7.637 ± 0.006, -7.643 ± 0.003, -7.636 ± 0.006, -7.634 ± 0.004) (P < 0.05). Single factor linear correlation analysis showed that there was a significant negative correlation between RSTN expression and QUICKI (-0.983, P < 0.05) in the traumatic group. The increase in RSTN expression in the subcutaneous adipose tissue of rats with traumatic brain injury is likely related to the indexes of glycometabolism, including serum insulin, fasting blood glucose, and QUICKI. Our results lead us to conclude that RSTN may play an important role in the process of insulin resistance in rats with traumatic brain injury.

Characterization of microsatellite markers and their correlations with growth traits in Mandarin fish (Siniperca chuatsi).

Mandarin fish (Siniperca chuatsi) is a traditionally cultured freshwater fish with high commercial value in China. To facilitate marker-assisted selection for genetic improvement of this species, 100 microsatellite markers identified in previous studies were characterized in the 25 largest and 25 smallest individuals. Twenty polymorphic loci were used to genotype 200 individuals, and the associations between their genotypes and growth traits were examined. We found that 9 genotypes at 8 loci (SC-10, Sin 135, Sin 166, AP 34-23, AP 38-11, AP 37-22, AP 37-08, and AP 37-37) were positively correlated with growth traits (body weight, body length, body height) in the mandarin fish population. The average of observed and expected heterozygosities were 0.71 and 0.59, respectively, and the average polymorphism information content value was 0.54, indicating that the population had high genetic diversity. The markers developed in this study are useful for selection of genetic breeding in this species and its related species.

Microsatellite DNA markers and their correlation with growth traits in mandarin fish (Siniperca chuatsi).

The mandarin fish (Siniperca chuatsi) is a traditionally cultured freshwater fish with high commercial value in China. To facilitate marker-assisted selection in genetic improvement of this species, 120 microsatellite markers from the literature were characterized in the 25 largest and 25 smallest individuals. Eighteen polymorphic loci were then used to genotype 200 individuals, and the associations between their genotypes and growth traits were examined. We found that eight genotypes of six loci (AP 37-06, AP 37-11, AP 37-16, AP 37-48, AP 38-32, and AP 39-05) were positively correlated with growth traits (body weight, length, and height) in the mandarin fish population. The average observed and expected heterozygosities were 0.68 and 0.59, respectively, and the average PIC value was 0.50, indicating a population with high genetic diversity. Therefore, these markers could be useful for assisted selection in genetic breeding of this species and its related species.

Analysis of DNA methylation patterns and levels in maize hybrids and their parents.

Heterosis is the superior performance of heterozygous individuals and has been widely exploited in plant breeding, although the underlying regulatory mechanisms still remain largely elusive. To understand the molecular basis of heterosis in maize, in this study, roots and leaves at the seedling stage and embryos and endosperm tissues 15 days after fertilization of 2 elite hybrids and their parental lines were used to estimate the levels and patterns of cytosine methylation by the methylation-sensitive amplification polymorphism method. The relative total methylation levels were lower in all the tissues of all hybrids than their corresponding mid-parent values, and the number of demethylation events was higher in the hybrids. These results implied that the decreasing trend and demethylation in hybrids relative to their parents may enable the derepression and possibly expression of many genes that were associated with the phenotypic variation in hybrids. To further analyze the observed methylation pattern changes, a total of 63 differentially displayed DNA fragments were successfully sequenced. Basic Local Alignment Search Tool analysis showed that 11 fragments shared similarity with known functional proteins in maize or other plant species, including metabolism, transposon/retrotransposon, development, stress response, and signal transduction, which indicated that these genes might play a significant role in maize hybrid vigor.

Cloning and characterization of a ß-amyrin synthase gene from the medicinal tree Aralia elata (Araliaceae).

Aralia elata is an important medicinal plant in China; it produces large amounts of oleanane type triterpene saponins. A full-length cDNA encoding ß-amyrin synthase (designated as AeAS) was isolated from young leaves of A. elata by reverse transcription-PCR. The full-length cDNA of AeAS was found to have a 2292-bp open reading frame, encoding a protein with 763 amino acid residues. The deduced amino acid sequence of AeAS showed the highest identity (97%) to Panax ginseng ß-amyrin synthase. When AeAS cDNA was expressed in Escherichia coli, an 87.8-kDa recombinant protein was detected by SDS-PAGE and Western blotting. The sequence was also heterologously expressed in the yeast Pichia pastoris, and production of ß-amyrin was detected by HPLC. Tissue expression pattern analysis by real-time reverse transcription-PCR revealed that AeAS is strongly expressed in leaves and stems, and weakly expressed in roots and flowers.

Proteomic study of Tianxiangdan intervention in rats with myocardial ischemia.

The present study aimed to screen out the proteins with significantly differential expression through the proteomics study of Tianxiangdan intervention in rats with myocardial ischemia as well as elucidate the mechanism of the intervention. In total, 54 Wistar rats (male, 6-8 weeks old) were randomly divided into the blank group, sham operation group, and model group, with 6 rats in each, together with the model + low dose group, model + medium-dose group, and model + high dose group, with 12 rats in each. Upon successful construction of the ischemic model, low, medium, and high doses, respectively, of Tianxiangdan were administered in the groups. The rat model of coronary heart disease (CHD) with myocardial ischemia was prepared by ligating the coronary artery. The tandem mass tag-labeled quantitative proteomics technology was adopted to observe the differentially expressed proteins in the myocardium of the model rats under the action of Tianxiangdan to find the target proteins for the treatment of myocardial ischemia in CHD. A total of 3122 proteins were identified. Combined with the references, tropomyosin alpha-3 chain (TPM3), protein kinase C delta (PRKCD), myosin heavy chain 10 (MYH10), MYH6, G protein subunit alpha i2 (GNAI2), and other proteins were screened out. Western blotting was adopted for the proteomics validation, and it was found that compared with the sham operation group, the expression levels of the GNAI2, TPM3, and MYH10 proteins were upregulated in the myocardial ischemia model group but downregulated after the administration of Tianxiangdan; the differences were statistically significant (p<0.05). We conclude that Tianxiangdan could improve myocardial ischemia by downregulating the proteins, including GNAI2, TPM3, and MYH10, which might be potential targets of Tianxiangdan in the treatment of myocardial infarction.

[Clinical analysis of 7 children infected with Talaromyces marneffei].

Objective: To investigate the clinical manifestations, treatments, and prognosis of pediatric patients with Talaromyces marneffei infection. Methods: In this retrospective study, 7 children diagnosed with Talaromyces marneffei infection in Shenzhen Children's Hospital from July 2017 to October 2021 were recruited. The clinical features, radiology, pathogen detection, immunological evaluation, treatments, and prognosis were analyzed. Results: In 7 cases, 5 were male, 2 were females. The age was from 0.75 to 8.75 years. The main clinical manifestations were fever in 7 cases, cough in 6 cases, malnutrition in 4 cases, papules in 2 cases and medical history of recurrent infection in 3 cases. Physical examination showed that all 7 patients had hepatosplenomegaly, 4 had superficial lymphadenopathy. Laboratory examination showed that 6 cases had decreased hemoglobin and 3 cases had decreased platelet. Chest CT showed that 4 cases had patchy shadows, pleural effusion, mediastinal or axillary lymph node enlargement, 3 had nodular shadows and 2 had cavities. The positive ratio of Talaromyces marneffei culture was 2/2 with tissue samples, 4/5 with bone marrow. The positive ratio was 3/4 by metagenomic next generation sequencing. The fungus was detected in 3 cases by smear microscopy of bone marrow and (or) peripheral blood. All patients were negative for human immunodeficiency virus by the immune function assay. However, 5 cases were confirmed as primary immunodeficiency disease, including 2 cases with high IgM syndrome, 2 with STAT1 gene variation, and the last with severe combined immunodeficiency (IL2RG gene variation). Exclude 1 case which gave up treatment due to acute intracranial infection, and the other patients received effective treatments along with amphotericin B, voriconazole, and itraconazole alone or in combination. Two cases relapsed after medication withdrawal, but 1 case got complete rehabilitation after hematopoietic stem cell transplantation. Conclusions: The clinical manifestations involve multisystem, the common charateristics are fever and cough. The chest CT imaging manifestations are diverse, it should be considered in differentiating tuberculosis. The amphotericin B, voriconazole and itraconazole are effective, but it will easily relapse when withdrawing those antifungal agents.

MicroRNA-488 inhibits ovarian cancer cell metastasis through regulating CCNG1 and p53 expression.

OBJECTIVE: The roles of microRNAs (miRNAs) have been widely exploited in cancer. MiRNAs have become a potential breakthrough in cancer diagnosis and treatment. Here, the regulatory mechanism of microRNA-488 (miR-488) was investigated in ovarian cancer (OC). PATIENTS AND METHODS: The expression levels of miR-488 and CCNG1 (Cyclin G1) were detected by quantitative Real Time-Polymerase Chain Reaction (qRT-PCR) and Western blot assays. Transwell assay and epithelial-mesenchymal transition (EMT) markers were used to clarify the effect of miR-488 on cell metastasis. The dual-luciferase reporter assay was used to verify the relation between miR-488 and CCNG1. RESULTS: The expression of miR-488 was reduced in OC, which was associated with poor clinical outcomes and prognosis in OC patients. MiR-488 inhibited cell metastasis in OC by blocking EMT and promoting tumor suppressor p53 expression. In addition, CCNG1 was confirmed as a direct target of miR-488. Upregulation of CCNG1 impaired the inhibitory effect of miR-488 in OC. CONCLUSIONS: MiR-488 serves as a tumor inhibitor in OC by suppressing cell metastasis, indicating that miR-488 has a great potential in the diagnosis and treatment of OC.

[Cilia ultrastructural and gene variation of primary ciliary dyskinesia: report of three cases and literatures review].

Objective: To analyze the clinical manifestations, cilia ultrastructure and gene variations of primary ciliary dyskinesia (PCD). Methods: Analysis of three cases diagnosed as PCD by transmission electron microscopy of the endobronchial biopsy material in Division of Pediatric Pulmonology of Shandong Provincial Hospital between 2013 and 2016. Target gene sequence capture and next generation sequencing were used to analyze the gene. Related literatures on gene variation of PCD in Chinese were reviewed from Online Mendelian Inheritance in Man, Human Gene Mutation Database, PubMed and CNKI up to July 2017 by using search terms of "PCD" , "gene" , "Chinese". Results: There were one male and two females aged from 10 to 11 years. The common symptoms included recurrent respiratory infection, sinusitis and bronchiectasis. Two of them had situs inversus. Case 1 had lack of outer and inner dynein arms with compound heterozygous mutation of LRRC6. Case 2 had outer and inner dynein arms defects with heterozygous mutations of DNAH5 and DNAH11. Case 3 had abnormality in microtubule and inner dynein arms with homozygous mutation of CCDC39. All the variations mentioned above have not been reported before. Twelve cases have been reported about gene variations in PCD in Chinese from eight reports. All these patients had recurrent respiratory infection starting soon after birth, rhinosinusitis, and bronchiectasis. Nine of them had dextrocardia. Four cases have taken an effective nasal (or bronchial) mucosal biopsy. 1 case had inner and outer dynein arms defects. One case had inner dynein arms and radial spokes defects. One case had microtubule and central pair defects. And 1 case had normal cilia ultrastructure. Eight kinds of gene variations were found. Three cases had gene variations of DNAH5. 2 cases had gene variations of DYX1C1. 2 cases had gene variations of CCNO. There was 1 case with gene variations of CCDC39, CCDC40, HYDIN, ARMC4 and DNAI1 separately. Conclusions: Recurrent respiratory infection starting soon after birth, rhinosinusitis, and bronchiectasis are the common symptoms of PCD. Eleven of fifteen Chinese PCD patients with positive gene mutations were Kartagener syndrome. Cilia ultrastructure showed defects of inner and outer dynein arms, radial spokes, microtubule and central pair. Ten kinds of gene variations were found: DNAH5, DYX1C1, CCNO, CCDC39, CCDC40, HYDIN, ARMC4, DNAI1, LRRC6、DNAH11.