[Clinicopathological features of hepatic fibrinogen storage disease in children].

Objective: To investigate the clinicopathological and molecular characteristics of hepatic fibrinogen storage disease (FSD) in children. Methods: The clinical, histopathologic, immunophenotypic, ultrastructural and gene sequencing data of 4 FSD cases were collected from September 2019 to January 2021 in the Children's Hospital of Fudan University, Shanghai, China. Retrospective analysis and literature review were conducted. Results: There were 4 cases of FSD, 3 males and 1 female, aged 3 years and 3 months to 6 years (median age, 3 years and 4 months). The clinical manifestations were abnormal liver function and abnormal blood coagulation function, for which 2 cases had family genetic history. Liver biopsies revealed that, besides liver steatosis, fibrosis and inflammation, there were single or multiple eosinophilic inclusion bodies of various sizes and surrounding transparent pale halo in hepatocytes. Immunohistochemistry showed that the inclusion bodies were positive for anti-fibrinogen. Under the electron microscope, they corresponded to the dilated cisternae of the rough endoplasmic reticulum, which were occupied by compactly packed tubular structures and arranged into a fingerprint-like pattern with curved bundles. Gene sequencing revealed that the 2 cases of FGG mutation were located in exon 8 c.1106A>G (p.His369Arg) and c.905T>C (p.Leu302Pro), and 1 case was located in exon 9 c.1201C>T (p.Arg401Trp). No pathogenic variant was detected in the other case. Conclusions: FSD is a rare genetic metabolic disease and clinically manifests as abnormal liver function with hypofibrinogenemia. In the background of liver steatosis, fibrosis and inflammation, there are eosinophilic inclusions with pale halo in the hepatocytic cytoplasm, which can be identified by anti-fibrinogen immunohistochemical staining. The fingerprint-like structures under electron microscope are helpful for the diagnosis, while FGG sequencing detects the pathogenic mutation of exon 8 or 9 that can clearly explain the phenotype. However, the diagnosis of FSD cannot be completely ruled out if the relevant mutations are not detected.

[GPCR48 promotes invasion and metastasis by inducing epithelial-mesenchymal transition in hepatocellular carcinoma cells].

Objective: To observe the G protein-coupled receptor 48 (GPCR48) expression in hepatocellular carcinoma (HCC) cell lines with different metastatic potential and its characteristics effect on the invasion and metastasis of Huh7 hepatoma cells via epithelial-mesenchymal transition (EMT). Methods: Western blot was used to detect the protein expression level of GPCR48 in HCC cells with different metastatic potential. The lentivirus vector expressing GPCR48 gene was constructed. GPCR48 was overexpressed in Huh7 hepatoma cells. The GPCR48 overexpression level was detected by real-time PCR and Western blot. Transwell invasion and migration assay was used to detect the Huh7 hepatoma cells invasion and migration ability in the Control, Mock and GPCR48 overexpression group. Real-time PCR and Western blot were used to detect Huh7 hepatoma cells mRNA and protein expression levels of the EMT related markers (E-cadherin, N-cadherin, vimentin, and γ catenin) in the Control, Mock and GPCR48 overexpression groups, respectively. Analysis of variance was used to compare the differences between data sets. Results: GPCR48 protein expression level in metastatic HCC cell lines was significantly higher than non-metastatic HCC cell lines (P < 0.05). The lentivirus vector expressing the GPCR48 gene had effectively transfected the Huh7 hepatoma cells and stably expressed the GPCR48mRNA and protein. Compared with the Mock and the Control group, Huh7 hepatoma cells invasion and migration ability in the GPCR48 overexpression group was significantly enhanced (F≥5.54, P < 0.05), and the mRNA and protein expression levels of epithelial phenotypic markers E-cadherin and γ-catenin were decreased (P < 0.05). The mRNA and protein expression levels of the mesenchymal phenotypic markers N-cadherin and Vimentin were increased (P < 0.05), indicating that EMT changes occurred in Huh7 hepatoma cells had overexpressed GPCR48. Conclusion: GPCR48 expression level is positively correlated with the metastatic potential of HCC cells. GPCR48 overexpression can down-regulate the expression of epithelial phenotypic markers and up-regulate the expression of mesenchymal phenotypic markers, and induce EMT changes in HCC cells, thus promoting HCC cells invasion and migration.

Brain-derived neurotrophic factor Val66Met genotype and ovarian steroids interactively modulate working memory-related hippocampal function in women: a multimodal neuroimaging study.

Preclinical evidence suggests that the actions of ovarian steroid hormones and brain-derived neurotrophic factor (BDNF) are highly convergent on brain function. Studies in humanized mice document an interaction between estrus cycle-related changes in estradiol secretion and BDNF Val66Met genotype on measures of hippocampal function and anxiety-like behavior. We believe our multimodal imaging data provide the first demonstration in women that the effects of the BDNF Val/Met polymorphism on hippocampal function are selectively modulated by estradiol. In a 6-month pharmacological hormone manipulation protocol, healthy, regularly menstruating, asymptomatic women completed positron emission tomography (PET) and functional magnetic resonance imaging (fMRI) scans while performing the n-back working memory task during three hormone conditions: ovarian suppression induced by the gonadotropin-releasing hormone agonist, leuprolide acetate; leuprolide plus estradiol; and leuprolide plus progesterone. For each of the three hormone conditions, a discovery data set was obtained with oxygen-15 water regional cerebral blood flow PET in 39 healthy women genotyped for BDNF Val66Met, and a confirmatory data set was obtained with fMRI in 27 women. Our results, in close agreement across the two imaging platforms, demonstrate an ovarian hormone-by-BDNF interaction on working memory-related hippocampal function (PET: F2,37=9.11, P=0.00026 uncorrected, P=0.05, familywise error corrected with small volume correction; fMRI: F2,25=5.43, P=0.01, uncorrected) that reflects differential hippocampal recruitment in Met carriers but only in the presence of estradiol. These findings have clinical relevance for understanding the neurobiological basis of individual differences in the cognitive and behavioral effects of ovarian steroids in women, and may provide a neurogenetic framework for understanding neuropsychiatric disorders related to reproductive hormones as well as illnesses with sex differences in disease expression and course.

[Clinical features, diagnosis and treatment of intraductal papillary neoplasm of the bile duct].

Objective: To study the clinicopathologic features of intraductal papillary neoplasm of the bile duct(IPNB) and to analyze the diagnostic and therapeutic patterns. Methods: The data of 46 patients with IPNB undergoing surgery in Department of Hepatobiliary and Pancreatic Surgery, the Second Affiliated Hospital of Zhejiang University School of Medicine from January 2013 to November 2017 were retrospectively analyzed.There were 23 males and 23 females with age of (64±8)years.Patients were followed up by clinics and telephone inquiry.Categorical data were compared with χ(2) test or Fisher's exact test. Results: Abdominal pain(in 31 patients), fever (in 15 patients) and jaundice (in 11 patients) were the most common symptoms.Twenty-five patients were accompanied with cholangiolithiasis and 25 were accompanied with liver atrophy.Preoperative laboratory examination was mainly manifested as the abnormal liver function caused by biliary obstruction.Typical imaging findings included bile duct dilation (in 45 patients) and mass within bile duct (in 22 patients). All the patients were diagnosed as IPNB histopathologically.Among them, high-grade intraepithelial neoplasia and related adenocarcinoma were more common in mucus-hypersecretion IPNB ((13/15 vs. 51.6%(16/31))(χ(2)=5.331, P=0.021). Hepatectomy was performed in 25 patients, hepatectomy combined with biliary resection and reconstruction in 12 cases, biliary resection and reconstruction in 3 cases, pancreatoduodenectomy in 3 cases, hepatopancreaticoduodenectomy in 1 case, liver transplantation in 1 case and radiofrequency ablation in 1 case.Forty-one patients were followed up with a median of 30 (12, 41) months.Seven patients suffered recurrence and 6 died. Conclusion: IPNB is a rare disease with limited knowledge currently.Images are the main diagnositc means and surgery is the first choice.

Brain-derived neurotrophic factor (BDNF) Val(66)Met polymorphism differentially predicts hippocampal function in medication-free patients with schizophrenia.

A Val(66)Met single-nucleotide polymorphism (SNP) in the brain-derived neurotrophic factor (BDNF) gene impairs activity-dependent BDNF release in cultured hippocampal neurons and predicts impaired memory and exaggerated basal hippocampal activity in healthy humans. Several clinical genetic association studies along with multi-modal evidence for hippocampal dysfunction in schizophrenia indirectly suggest a relationship between schizophrenia and genetically determined BDNF function in the hippocampus. To directly test this hypothesized relationship, we studied 47 medication-free patients with schizophrenia or schizoaffective disorder and 74 healthy comparison individuals with genotyping for the Val(66)Met SNP and [(15)O]H(2)O positron emission tomography (PET) to measure resting and working memory-related hippocampal regional cerebral blood flow (rCBF). In patients, harboring a Met allele was associated with significantly less hippocampal rCBF. This finding was opposite to the genotype effect seen in healthy participants, resulting in a significant diagnosis-by-genotype interaction. Exploratory analyses of interregional resting rCBF covariation revealed a specific and significant diagnosis-by-genotype interaction effect on hippocampal-prefrontal coupling. A diagnosis-by-genotype interaction was also found for working memory-related hippocampal rCBF change, which was uniquely attenuated in Met allele-carrying patients. Thus, both task-independent and task-dependent hippocampal neurophysiology accommodates a Met allelic background differently in patients with schizophrenia than in control subjects. Potentially consistent with the hypothesis that cellular sequelae of the BDNF Val(66)Met SNP interface with aspects of schizophrenic hippocampal and frontotemporal dysfunction, these results warrant future investigation to understand the contributions of unique patient trait or state variables to these robust interactions.

Protective effect of allicin on ischemia-reperfusion injury caused by testicular torsion-detorsion in rats.

OBJECTIVE: Testicular ischemia-reperfusion induced by testicular torsion-detorsion increases the level of reactive oxygen species, leading to testicular damage. Allicin, one of the most active ingredients in garlic, is a significant exogenous antioxidant. In the research, the efficacy of allicin in treating testicular ischemia-reperfusion injury was assessed. MATERIALS AND METHODS: The study included sixty Sprague-Dawley male rats. Three groups with 20 rats per group were created as follows: control group, testicular ischemia/reperfusion-induced group, and testicular ischemia-reperfusion plus treatment with allicin group. The control group underwent a sham operation of the left testis without other interventions. In the testicular ischemia/reperfusion-induced group, rat left testis was subjected to 720° torsion for two hours and then detorsion. In the allicin-treated group, in addition to testicular ischemia-reperfusion, 50 mg/kg of allicin was injected intraperitoneally, starting immediately following detorsion. Testicular tissue samples were obtained to measure the protein expression of xanthine oxidase, which is a major source of reactive oxygen species formation, malondialdehyde level (a reliable marker of reactive oxygen species), and testicular spermatogenic function. RESULTS: Testicular ischemia-reperfusion significantly increased the expression of xanthine oxidase and malondialdehyde levels in ipsilateral testes while reducing testicular spermatogenic function. The expression of xanthine oxidase and malondialdehyde levels were significantly lower in ipsilateral testes, whereas testicular spermatogenic function in the allicin-treated group was significantly higher compared with those in the testicular ischemia-reperfusion group. CONCLUSIONS: Our findings indicate that allicin administration improves ischemia/reperfusion-induced testicular damage by limiting reactive oxygen species generation via inhibition of xanthine oxidase expression.

Elastin peptide concentration in human serum: variation with antibodies and elastin peptides used for the enzyme-linked immunosorbent assay.

Discrepancies exist between the reported values for the mean elastin peptide (EP) concentration in human sera. In order to understand these discrepancies, several EP preparations were obtained in vitro and monoclonal and polyclonal antibodies were produced against them. These different EP preparations and antibodies were used in an enzyme-linked immunosorbent assay (ELISA) to study cross-reactivity between EP preparations and to quantitate EP concentration in human sera. The method of purification of elastin, the method of hydrolysis of elastin and the molecular weight of EP influence their reactivity with antibodies and the results of EP measurements in human sera. However, there is a good correlation between EP measurements carried out in several human sera with the different EP preparations and different antibodies. Although absolute values of the EP concentrations varied with the EP preparation and antibodies used for the ELISA, the variations of this EP concentration measured from one human serum to another are significant.

Ultraviolet radiation and reactive oxygen generation as inducers of keratinocyte apoptosis: protective role of tea polyphenols.

Ultraviolet A (UVA) radiation produces serious damage to skin, especially to dermis, but its damage to epidermis and responsible mechanisms are not fully understood. Studies were thus undertaken to investigate the effects of UVA or reactive oxygen species (ROS) on lipid peroxidation, cell cycle, and apoptosis in primary cultured rat keratinocytes and to determine the possible protective effects of tea polyphenols (TPP). UVA or ROS increased the release of plasma enzyme lactate dehydrogenase (LDH), and increased lipid peroxidation production (malondialdehyde, MDA), but decreased the activity of glutathione peroxidase (GSH-Px), indicating that UVA or ROS were cytostatic and peroxidizing to keratinocytes. TPP stabilized and protected cell membranes from ROS or UVA by inhibiting the release of LDH, lowering MDA levels, and increasing GSH-Px activity. Flow cytometry (FCM) analysis revealed that UVA or ROS decreased the proliferative index (PI); hence the cell growth was blocked in the S/G2 phase, with an increase in the percentage of apoptosis in primary keratinocytes. TPP modified the UVA or ROS-induced changes in PI and apoptosis. TPP may be useful to protect keratinocytes from UVA irradiation. In summary, these data demonstrated that UVA damage to skin keratinocytes in vitro was similar to that for ROS and that TPP protects against UVA-induced cytotoxicity by inhibiting lipid peroxidation and apoptosis.

The effects on cell growth of tea polyphenols acting as a strong anti-peroxidatant and an inhibitor of apoptosis in primary cultured rat skin cells.

Studies during the past few years have indicated an inhibitory effect of green tea or tea polyphenols on tumorigenesis in animal and even in human. The purpose of this study was to observe the possible effects of tea polyphenols on skin cell growth and on apoptosis in rat primary cultured keratinocytes and fibroblasts. The release of a cell plasma enzyme (LDH), lipid peroxidation products (MDA production), and GSH-Px (glutathione peroxidase) into the medium in cultured cells was determined after treatment with tea polyphenols in a primary culture of skin cells. The percentage of cells in each cell cycle phase and in apoptosis were assayed by flow cytometry (FCM). Tea polyphenols may have a beneficial effect on skin cells at concentrations from 0.05% to 0.1%, showing a dose-dependent decrease in LDH, MDA (malondialdehyde) production, and a significant dose-dependent increase in GSH-Px and cell number. These effects were more obvious after exposure for 24 h than after 12 h. The results indicate that tea polyphenols may stabilize and protect the cell membrane against the release of cell plasma enzyme LDH, and its anti-peroxidation effect is also important for cell growth. FCM analysis revealed that treatment with 0.01% to 0.1% tea polyphenols decreased the percentage of cells in the G1/G0 (quiescent) phase from 81.32% to 74.38%, and increased the percentage of cells in S and G2/M phase from 9.87% to 15.26%, and from 6.51% to 10.36%, respectively. Tea polyphenols also increased the value of PI (proliferation index) from 18.17 to 25.62. At the same time it decreased the percentage of apoptosis from 27.10% to 17.97%, which indicates that green tea stimulates cell growth and inhibits the occurrence of apoptosis. Our results indicate that tea polyphenols are effective anti-oxidants and also inhibit apoptosis, which may improve the proliferative capacity of primary skin cells in vitro.

[Monoclonal antibodies against gastric cancer and their selective reaction on various tissues].

Spleen cells from Balb/c mice immunized in sequence with five human gastric cancer cell lines were fused with murine myeloma cell line SP2/0. Hybridomas 3F4, 3G9 and 3H11 secreting monoclonal antibodies (mAb) against gastric cancer were obtained through selective culture and screening. These mAb produced by the immunization procedure have good selectivity and high positive rate in reaction on gastric cancer. The positive rate of reaction on gastric cancer cells and tissues could reach 5/5 and 84.8-93.5%, respectively, whereas there was almost no positive reaction on normal cells and tissues As there was no correlation between the positive reaction of gastric cancer and their histopathologic typing, and the cross reaction of mAb with other tumors and fetal gastrointestinal tissues was quite high, the corresponding antigens of these mAb were considered as extensive oncofetal antigens.

[Monoclonal antibody A12 against gastric cancer produced in immuno-reconstituted nude mice].

A monoclonal antibody (mAb) A12 against gastric cancer was prepared in immuno-reconstituted nude mice from human ductal adenocarcinoma of stomach, Sy86B. mAb A12 could react with the majority of gastric cancer tissues (24/27-88.9%) but only cross react with a few normal tissues tested. The corresponding antigen of mAb A12 (antigen A12) was expressed at higher levels and usually in more than 50% of the cancer cells. mAb A12 may be a valid preparation in targeting therapy of gastric cancer. Preliminary analysis of antigen A12 showed that it is a oncofetal antigen probably of glycolipid or glycoprotein in nature.

[Immunoelectron microscopic study on the localization of monoclonal antibodies on gastric cancer cells].

The localization of three monoclonal antibodies (mAb) against gastric cancer was studied on two human gastric cancer cell lines by immunoelectron microscopic technique. It was shown that the corresponding antigens of mAb 3G9 and 3H11 were distributed on the microvilli (M) and non-microvillous (NM) plasma membrane of target cells, with varying M to NM ratios depending on the mAbs and target cells used. However, the corresponding antigens of mAb, PD4 was only localized on the surface of round or finger-like bulges of target cells and never on the microvilli and non-microvillous plasma membrane. Since the nature and function of these tumor antigens are yet to be identified, the implication of the different distributions of these tumor antigens remains to be claifated.

[Gastric cancer-associated antigen P42].

The corresponding antigen to monoclonal antibody PCI against gastric cancer cells was demonstrated as a protein sensitive to heating and easily degraded by proteinase. By western blotting, it was shown that the molecular weight of the antigen was 42 KD and the antigen was named P42. P42 could be shed into serum-free culture medium of the target cells and its activity be assayed in sera of the gastric cancer patients. Using inhibition test, the levels of P42-like substance in sera were measured. The results indicated that this level in the gastric cancer patients (26.53 +/- 23.11) was markedly higher than that in patients with benign gastric diseases (4.66 +/- 2.67) and controls (2.57 +/- 2.67). It is suggested that P42, a novel gastric cancer-associated antigen, be useful in the mass screening and supplementary in diagnosing gastric cancer.