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Background and aims Familial hypercholesterolemia (FH) is an autosomal dominant disease with widespread global prevalence that partially accounts for the high prevalence of premature coronary heart disease. Although the majority of research on FH has focused on single heterozygous LDLR mutations, there have been limited reports of double LDLR mutations on the same chromosome. The aim of this study was to gain insight into the clinical consequences of the presence of multiple mutations in the LDLR gene. Methods DNA from two clinical homozygous FH patients and their relatives was analysed using targeted exome sequencing and DNA resequencing. Functional characterization of novel variants was performed by Western blot, flow cytometry and confocal microscopy. Results Proband 1 carried p.Q12X, NTDA (p.N276T and c.892delA) mutations in LDLR , and Proband 2 carried c.971delG, GSDN (p.G77S + D601N). Results showed that p.Q12X, c.892delA, and c.971delG are non-functional LDLR variants. Conversely, N276T and G77S are non-pathogenic variants. Interestingly, while D601N alone only slightly diminishes LDLR activity, its co-presence with the non pathogenic p.G77S mutation results in a more strongly pathogenic variant with LDLR activity reduced by 40%. One of the double mutants, NTDA, is as non functional as c.892delA alone. The other double mutant, GSDN, is more severe than either of the component single mutants. Conclusions An early gene screening and laboratory functional verification of LDLR activity is of vital importance to enable a definite FH diagnosis. Functional verification is also necessary for prenatal and postnatal care in patients with FH.

// Dong-Hua Gu 1, * , Jia-Hui Mao 3, * , Xiao-Dong Pan 1, 2, * , Hua Zhu 2 , Xinfeng Chen 2 , Bing Zheng 2 , Yuxi Shan 1 1 The Department of Urology, The Second Affiliated Hospital of Soochow University, Suzhou, China 2 The Department of Urology, The Second Affiliated Hospital of Nantong University, Nantong, China 3 Department of Pathophysiology, Nantong University School of Medicine, Nantong, China * These authors contributed equally to this work Correspondence to: Bing Zheng, email: bingzhengnantong@163.com Yuxi Shan, email: drshanyuxisd@163.com Keywords: renal cell carcinoma (RCC), Grb2-associated binding 2 (Gab2), microRNA-302c-3p, Akt and cell proliferation Received: January 10, 2017 Accepted: February 02, 2017 Published: February 17, 2017 ABSTRACT The expression and biological function of Grb2-associated binding 2 (Gab2) in renal cell carcinoma (RCC) cells was tested here. We showed that Gab2 expression was significantly elevated in human RCC tissues and RCC cells. It was correlated with over-activation of Akt and downregulation of microRNA-302c-3p (“miR-302c-3p”), a putative Gab2-targeting microRNA. Knockdown of Gab2 inhibited Akt activation and 786-O RCC cell proliferation. Reversely, forced over-expression of Gab2 led to Akt hyper-activation to facilitate 786-O cell proliferation. Exogenous expression of miR-302c caused Gab2 downregulation, Akt inhibition and 786-O cell proliferation inhibition. On the other hand, miR-302c-3p depletion by expressing its anti-sense (“antagomiR-302c”) led to Gab2 upregulation, Akt activation and increased 786-O cell proliferation. Significantly, miR-302c-3p failed to affect the proliferation of 786-O cells with shRNA-depleted Gab2. Together, we suggest that miR-302c-3p depletion in human RCC cells leads to Gab2 over-expression, Akt hyper-activation and cell proliferation.

We described a method for quantification of IgG (IgG1, IgG2, IgG3 and IgG4) in a small amount of human blood plasma using high-resolution quadrupole-Orbitrap mass spectrometry (HR-Q-Orbitrap-MS). For screening surrogate peptides of targeted IgGs, samples were prepared by pellet digestion and identified by data-independent acquisition of Orbitrap MS. The quantification was performed by parallel reaction monitoring (PRM). The remarkable sensitivity and selectivity of PRM enable the detection of low abundance IgGs, or a little plasma. The assay linearity for external standard IgG proteins was obtained from 5 to 1000 μg L−1 with a mean coefficient of determination (R2) higher than 0.99. The inter-day (n = 6) and intra-day (n = 6) precisions obtained were from 3.6% to 11.5%. This report represents a comprehensive study so far of the use of Q-Orbitrap-MS for the identification and quantitation of IgGs in human blood plasma.

A method for shrimp muscle identification in complex foods is required to safeguard the shrimp-allergic population. This study described a method for authentication of shrimp in complex foodstuffs (fish balls) by liquid chromatography tandem QTOF mass spectrometry (UPLC-QTOF-MS). The proteins in shrimp muscle were extracted using a Tris–HCl solution and then digested using tryptic protease. The main allergen proteins, tropomyosin (TM) and arginine kinase (AK), were characterized using the ‘bottom up’ MS approach. After analysis of their peptide mass fingerprinting based on the UniProt database, two specific heat-stable peptides, ALSNAEGEVAALNR for TM and VSSTLSSLEGELK for AK, were screened as surrogate (signature) peptides. The detection limit, expressed as shrimp meat per kilogram of food, was 8 g kg−1 (usage of TM) or 5 g kg−1 (usage of AK). The developed method is suitable to screen potential addition of shrimp meat in foodstuffs by detection of allergen proteins.

We developed a LC-MS/MS method for quantification of human lactoferrin in breast milk based on tryptic peptides and a synthetic isotopic peptide standard. The signature peptides were obtained from tryptic breast milk. They were screened by computational prediction using Biolynx software, and confirmed by database searching after analysis using liquid chromatography-quadrupole-time-of-flight tandem mass spectrometry (LC-Q-TOF-MS). The winged isotopic-labeled signature peptide was used as an internal standard to compensate the matrix effect. The spiking recovery of human lactoferrin ranged from 92.1% to 97.5%, and the relative standard deviation (RSD) was 3.4–4.7%. The limit of detection (LOD) and limit of quantitation (LOQ) were 1 mg/100 g and 3 mg/100 g in tryptic breast milk, respectively. The present testing method was sensitive and selective, and was successfully applied to human breast milk at different lactation stages in Beijing, China. The data revealed that levels of secreted lactoferrin decreased with the extension of lactation.

A method has been developed for quantitative analysis of vitamin K1 in vegetables and fruits by LC-MS/MS. The method employed a neutral aluminum oxide column for sample clean-up. Considering matrix effects, an isotope standard (vitamin K1-D7) was added to correct matrix interferences, as well as pretreatment losses and instrument variability. Spiked recoveries in model matrices were 88.3–90.1% for spinach, 84–110% for lotus roots, and 108–115.6% for grapes. The relative standard deviations (RSDs) were 1.0–7.1% for intra-day and 2.8–8.8% for inter-day. The limit of detection (LOD) and limit of quantification (LOQ) were 0.001 mg kg−1 and 0.004 mg kg−1, respectively. The method is sensitive and reliable, which is successfully applied to different vegetable and fruit matrices. Vitamin K1 in fruit samples ranged from 0.008 mg kg−1 to 0.401 mg kg−1, and in vegetable samples from 0.0032 mg kg−1 to 3.973 mg kg−1.

Apoplastic ascorbate (ASCapo) is an important contributor to the detoxification of ozone (O3). The objective of the study is to explore whether ASCapo is stimulated by elevated O3 concentrations. The detoxification of O3 by ASCapo was quantified in tobacco (Nicotiana L), soybean (Glycine max (L.) Merr.) and poplar (Populus L), which were exposed to charcoal-filtered air (CF) and elevated O3 treatments (E-O3). ASCapo in the three species were significantly increased by E-O3 compared with the values in the filtered treatment. For all three species, E-O3 significantly increased the malondialdehyde (MDA) content and decreased light-saturated rate of photosynthesis (Asat), suggesting that high O3 has induced injury/damage to plants. E-O3 significantly increased redox state in the apoplast (redox stateapo) for all species, whereas no effect on the apoplastic dehydroascorbate (DHAapo) was observed. In leaf tissues, E-O3 significantly enhanced reduced-ascorbate (ASC) and total ascorbate (ASC+DHA) in soybean and poplar, but significantly reduced these in tobacco, indicating different antioxidative capacity to the high O3 levels among the three species. Total antioxidant capacity in the apoplast (TACapo) was significantly increased by E-O3 in tobacco and poplar, but leaf tissue TAC was significantly enhanced only in tobacco. Leaf tissue superoxide anion (O2•-) in poplar and hydrogen peroxide (H2O2) in tobacco and soybean were significantly increased by E-O3. The diurnal variation of ASCapo, with maximum values occurring in the late morning and lower values experienced in the afternoon, appeared to play an important role in the harmful effects of O3 on tobacco, soybean and poplar.

Concern about metals and metalloids, especially heavy metals in seaweeds has risen due to potential health risk. This study investigated the distribution of 10 metals and metalloids in 295 dried seaweeds (brown and red) and estimated the possible health risk via hazard index (HI). Elements in seaweeds can be sequenced in descending order by mean values: Al > Mn > As > Cu > Cr > Ni > Cd > Se > Pb > Hg. The levels of Cd, Cu, Mn and Ni in red seaweeds were significantly higher than those in brown seaweeds (P r = 0.59, P

Chemotherapy-induced autophagy activation often contributes to cancer resistance. MiRNA-30a (miR-30a) is a potent inhibitor of autophagy by downregulating Beclin-1. In this study, we characterized the role of miR-30a in sorafenib-induced activity in renal cell carcinoma (RCC) cells. We found that expression of miR-30a was significantly downregulated in several human RCC tissues and in RCC cell lines. Accordingly, its targeted gene Beclin-1 was upregulated. Sorafenib activated autophagy in RCC cells (786-0 and A489 lines), evidenced by p62 degradation, Beclin-1/autophagy protein 5 (ATG-5) upregulation and light chain (LC)3B-I/-II conversion. Exogenously expressing miR-30a in 786-0 or A489 cells inhibited Beclin-1 expression and enhanced sorafenib-induced cytotoxicity. In contrast, knockdown of miR-30a by introducing antagomiR-30a increased Beclin-1 expression, and inhibited sorafenib-induced cytotoxicity against RCC cells. Autophagy inhibitors, including chloroquine, 3-methyaldenine or Bafliomycin A1, enhanced sorafenib activity, causing substantial cell apoptosis. Meanwhile, knockdown of Beclin-1 or ATG-5 by targeted siRNAs also increased sorafenib-induced cytotoxicity in above RCC cells. These findings indicate that dysregulation of miR-30a in RCC may interfere with the effectiveness of sorafenib-mediated apoptosis by an autophagy-dependent pathway, thus representing a novel potential therapeutic target for RCC.

Polyomavirus (BKV) and cytomegalovirus (CMV) are associated with renal graft failure. The aim was to establish a quantitative PCR method (Q-PCR) to detect BKV and CMV simultaneously. The conserved sequences of BKV and CMV were amplified and cloned into the plasmids as standards. The sensitivity, specificity and the precision of the assay were evaluated. Q-PCR was used to detect BKV and CMV DNA simultaneously in 480 blood samples of renal transplantation recipients. The sensitivity of the Q-PCR assay to detect BKV or CMV DNA reached 5×10(3)copies/mL. The use of control DNA verified that the assay could specifically detect the target DNA. The precision of the assay to quantify target DNA copies was acceptable (ICV 3.44% for BKV and 2.23% for CMV; differences between batches ICV 4.98% for BKV and 3.76% for CMV). In 480 samples, 130 samples (27.08%) were CMV DNA positive, which was significantly higher than the 64 BKV DNA positive samples (13.33%, p

Summary Background Baroreflex gain increase up closely to adult level during initial postnatal weeks, and any interruption within this period will increase the risk of cardiovascular problems in later of life span. We hypothesize that this short period after birth might be critical for postnatal development of vagal ganglion neurons (VGNs). Methods To evaluate neuroexcitability evidenced by discharge profiles and coordinate changes, ion currents were collected from identified A- and C-type VGNs at different developmental stages using whole-cell patch clamping. Results C-type VGNs underwent significant age-dependent transition from single action potential (AP) to repetitive discharge. The coordinate changes between TTX-S and TTX-R Na+ currents were also confirmed and well simulated by computer modeling. Although 4-AP or iberiotoxin age dependently increased firing frequency, AP duration was prolonged in an opposite fashion, which paralleled well with postnatal changes in 4-AP- and iberiotoxin-sensitive K+ current activity, whereas less developmental changes were verified in A-types. Conclusion These data demonstrate for the first time that the neuroexcitability of C-type VGNs increases significantly compared with A-types within initial postnatal weeks evidenced by AP discharge profiles and coordinate ion channel changes, which explain, at least in part, that initial postnatal weeks may be crucial for ontogenesis in visceral afferent reflex function.

Familial hypercholesterolemia (FH) is an autosomal dominant disorder. Although genetic testing is an important tool for detecting FH-causing mutations in patients, diagnostic methods for young patients with severe hypercholesterolemia are understudied. This study compares the target exome sequencing (TES) technique with the DNA resequencing array technique on young patients with severe hypercholesterolemia. A total of 20 unrelated patients (mean age 14.8 years) with total cholesterol > 10 mmol/L were included. 12 patient samples were processed by DNA resequencing array, 14 patient samples were processed by TES, and 6 patient samples were processed by both methods. Functional characterization of novel mutations was performed by flow cytometry. The mutation detection rate (MDR) of DNA resequencing array was 75%, while the MDR of TES was 100%. A total of 27 different mutations in the LDLR were identified, including 3 novel mutations and 8 mutations with previously unknown pathogenicity. Functional characterization of c.673delA, c.1363delC, p.Leu575Phe and p.Leu582Phe variants found that all of them are pathogenic. Additionally, 7 patients were diagnosed with Heterozygous FH (HeFH) in which lipid levels were significantly higher than common HeFH patients. This data indicates that TES is a very efficient tool for genetic diagnosis in young patients with severe hypercholesterolemia.

Objective. Postoperative fatigue syndrome (POFS) is a general and main complication after surgery. However, there is no stable and standardized animal model for POFS. The aim of the present study was to establish a rodent model of POFS by small intestinal resection, with POFS evaluated by acknowledged physical and behavioral methods. Material and Methods. Forty-two Sprague-Dawley rats were randomly divided into four groups according to the length of a "middle" small intestinal resection: 0% (sham group; i.e., laparotomy alone), 10%, 40% and 70% groups, with corresponding lengths of small intestinal resections. Following surgery, the general state of health was evaluated. Tail suspension test, open field test and Morris water maze test were used to evaluate the degree of POFS. Serum albumin, transferrin, prealbumin and fibronectin were measured to assess the nutritional status, and superoxide dismutase (SOD) and malondialdehyde (MDA) were also measured. Results. As compared with the other three groups, the 70% small intestinal resection group showed the worst general state of health, decreased strength of the tail suspension test and decreased score of Morris water maze test (p < 0.05) after operation. All rats in whom the small intestinal resection was done demonstrated a certain degree of malnutrition and behavior of depression, and the 70% resection group had the lowest levels of transferrin, prealbumin and fibronectin as compared with the other groups (p < 0.05), as well as decreased SOD and increased MDA in serum (p < 0.05). Conclusions. Resection of 70% of the small intestine resulted in typical characteristics of POFS. As this procedure is simple, stable and easily reproducible, it may serve as a model for research on POFS.

In China, about 1 million people are expected to suffer from familial hypercholesterolemia (FH), with a similar prevalence of FH to that in Caucasian populations. The mutations underlying FH in China are largely unknown because only a few studies have been conducted. In the present study, DNA sequencing analysis was used to scan the low-density lipoprotein receptor (LDLR) and ApoB100 genes in a Chinese family with clinically diagnosed FH. The results showed that the proband had abnormal patterns at nucleotide 517 of exon 4 due to a C/T heterozygosity, and at 1757 of exon 12 due to an A/C heterozygosity in the LDLR gene. Two mutations, C152R and S565X, were identified in the LDLR protein of the proband. DNA analysis of other family members showed that the two mutations should be located in different alleles of the proband. By flow cytometry analysis, the p.S565X mutant receptor displays reduced binding and internalization activity (16 and 19%, respectively) compared with the wild-type receptor. The proband is a compound heterozygote due to the C152R and S565X mutations, which are the possible molecular mechanisms of the etiology of FH in this family.

Background and aims Familial hypercholesterolemia (FH) is an autosomal dominant disorder of lipoprotein metabolism caused by mutations in the low-density lipoprotein receptor (LDL-R) gene, leading to elevated levels of cholesterol and an increased risk of coronary heart disease. In this article, from four homozygous FH phenotype probands we identified disease causing mutations and analyzed the relationship between genotype and phenotype. Methods and results DNA sequencing identified five LDL-R point mutations in four unrelated families. We found a novel homozygous mutation (C210R), a homozygous mutation at W462X, a compound heterozygous mutation of C122Y and T383I, and a G>A intron 3 splice site homozygous mutation. The functional alteration caused by the novel C210R mutation was confirmed by FACS analysis. Four probands have high low-density lipoprotein cholesterol (LDL-C) levels, ranging from 14.65 to 27.66 mmol/L. Their heterozygous parents had relatively low levels. B-mode ultrasound supplemented by Doppler was used to examine aortic/mitral valve structural alterations and carotid intima-media thickness (ITM) in all probands. The ITM values were between 1.2 and 2.3 mm, much higher than the normal value of Conclusion Our data demonstrated that all the probands were associated with severe hypercholesterolemia, thick carotid IMT and a low CFVR (coronary flow velocity reserve) value. The novel mutation (C120Y) is a disease causing mutation.

The characteristics of potential probotic Lactobacillus acidophilus NIT isolated from infant feces were evaluated in vitro. The strain was examined for resistance to pH 2–4 and 1–3% bile, adhesion to Caco-2 cells, and antimicrobial activities against enteric pathogenic bacteria. L. acidophilus NIT was shown tolerance property to bile, acid and strong antimicrobial activity against tested enteropathogens by the well-diffusion method. Furthermore, good adhesion and significant potential for decreasing adhesion of pathogens to Caco-2 cells were observed in this experiment. These results showed that L. acidophilus NIT may be useful for improving probiotic formulae with respect to protection against enteric pathogenic infection.

The purpose of this study was to clarify effects of selected oligosaccharides on concentrations of cecal short-chain fatty acids (SCFAs), total large bowel wet weight and wall weight, and cecal microbiota levels in mice. Mice were respectively given gavage of selected fructooligosaccharides (FOS), galactooligosaccharides (GOS), mannanoligosaccharides (MOS), and chitooligosaccharides (COS) [1000 mg/(kg body weight·d)]. Control group was given physiological saline solution. After 14 d treatment, SCFAs and lactate in mice cecum were significantly increased (P

Aims: Lactobacilli should resist stress environments in industry process and gastrointestinal tract before exerting their beneficial effects. To explore the possible stabilizers in probiotic products, prebiotic oligosaccharides were investigated. Methods and Results: We investigated the effect of four selected oligosaccharides on the survival of probiotic Lactobacillus plantarum and L. acidophilus to simulated stress conditions. It was found that the tolerance of lactobacilli to simulated artificial gastrointestinal juice, heat treatment and phenol solution was obviously enhanced in fructo-oligosaccharides (FOS) and xylo-oligosaccharides (XOS) group. In addition, chito-oligosaccharides (COS), manno-oligosaccharides (MOS) and glucose also had positive effect compared with control group (without sugar). Conclusions: Prebiotic oligosaccharides, especially XOS and FOS added in medium have protection function to lactobacilli in stress environments. The protection function of oligosaccharides may correlate with the bacteria growth, which was stimulated by these oligosaccharides. Significance and Impact of the Study: Prebiotic oligosaccharides may be used as stabilizers in probiotic products.

Aims: To characterize the antimicrobial and adhesion ability of candidate probiotic Clostridium butyricum CB2 for farmed fish in vitro. Methods and Results: The potential probiotic Cl. butyricum CB2 had been evaluated for its adhesion capacity and antagonistic effect against two fish pathogens Aeromonas hydrophila and Vibrio anguillarum by the intestinal cell model. In addition, the aggregation ability and antimicrobial property on agar plate were assayed. The results indicated that the candidate probiotic Cl. butyricum CB2 have strong adhesion property and a higher antagonistic activity to Aer. hydrophila and V. anguillarum both on agar plate and cell model. Clostridium butyricum showed a higher aggregation which might be the reasons for bacteria adhesion and antimicrobial activity. Conclusions: The strain Cl. butyricum CB2 could be used as potential probiotic to inhibit pathogens growth and prevent their colonization in fish intestinal tract. Significance and Impact of the Study: This study revealed the antimicrobial and adhesion characteristic of Cl. butyricum CB2 which was selected as the potential probiotic to farmed fish.

Familial hypercholesterolaemia (FH) is a serious genetic metabolic disease. We identified a specific family in which the proband had typical homozygous phenotype of FH, but couldn’t detect any mutations in usual pathogenic genes using traditional sequencing. This study is the first attempt to use whole exome sequencing (WES) to identify the pathogenic genes in Chinese FH. The routine examinations were performed on all parentage members and WES on 5 members. We used bioinformatics methods to splice and filter out the pathogenic gene. Finally, Sanger sequencing and cDNA sequencing were used to verify the candidate genes. Half of parentage members had got hypercholesterolaemia. WES identified LDLR IVS8[−10] as a candidate mutation from 222,267 variations. The Sanger sequencing showed proband had a homozygous mutation inherited from his parents and this loci were cosegregated with FH phenotype. The cDNA sequencing revealed that this mutations caused abnormal shearing. This mutation was first identified in Chinese patients and this homozygous mutation is a new genetic type of FH. This is the first time that WES was used in Chinese FH patients. We detected a novel genetic type of LDLR homozygous mutation. WES is powerful tools to identify specific FH families with potentially pathogenic gene mutations.

Oxymatrine is an alkaloid, which is derived from the traditional Chinese herb, Sophora flavescens Aiton. Oxymatrine has been shown to exhibit anti-inflammatory, antiviral, and anticancer properties. The present study aimed to investigate the anticancer effects of oxymatrine in human pros- tate cancer cells, and the underlying molecular mechanisms of these effects. An MTT assay demonstrated that oxymatrine significantly inhibited the proliferation of prostate cancer cells in a time‑ and dose‑dependent manner. In addition, flow cytometry and a terminal deoxynucleotidyl transferase-medi- ated dUTP-biotin nick end-labeling assay suggested that oxymatrine treatment may induce prostate cancer cell apop- tosis in a dose-dependent manner. Furthermore, western blot analysis demonstrated a significant increase in the expression of p53 and bax, and a significant decrease in that of Bcl-2, in prostrate cancer cells in a dose-dependent manner. In vivo analysis demonstrated that oxymatrine inhibited tumor growth following subcutaneous inoculation of prostate cancer cells into nude mice. The results of the present study suggested that the antitumor properties of oxymatrine, may be associ- ated with the inhibition of cell proliferation, and induction of apoptosis, via the regulation of apoptosis-associated gene expression. Therefore, the results may provide a novel approach for the development of prostate cancer therapy using oxymatrine, which is derived from the traditional Chinese herb, Sophora flavescens

Background Familial hypercholesterolemia (FH) is an autosomal dominant disease that primarily results from mutations in the low-density lipoprotein receptor (LDLR) gene. We investigated two unrelated Chinese FH patients using gene screening and functional analysis to reveal the pathogenicity and the mechanism by which these mutations cause FH. Methods First, the LDLR gene was sequenced in these patients. Then, mutant receptors were transfected into human embryo kidney 293(HEK-293) cells, and a confocal laser-scanning microscope was used to observe the localization of mutant proteins. Further, the expression and the internalization activity were analyzed by flow cytometry. Finally, LDLR protein expression and stability was detected by western blot. Results Two different LDLR class 2B mutations were detected in two patients. The C201F mutation is a known mutation. However, the G615V mutation is novel. Flow cytometry showed that the expression and internalization activity of the mutant LDLRs were reduced to 73.6% and 82.6% for G615V and 33.2% and 33.5% for C201F, respectively. Conclusions This study identified two LDLR mutations in Chinese patients with FH and analyzed the relationship between the genotype and phenotype of these patients. We found that these mutant LDLRs were defective in transport, which led to a reduction in cholesterol clearance. These results increase our understanding of the mutational spectrum of FH in the Chinese population.

Familial hypercholesterolemia is an autosomal dominant inherited disease characterized by elevated plasma low-density lipoprotein cholesterol (LDL-C). It is mainly caused by mutations of the low-density lipoprotein receptor (LDLR) gene. Currently, the methods of whole genome sequencing or whole exome sequencing for screening mutations in familial hypercholesterolemia are not applicable in China due to high cost. We performed targeted exome sequencing of 167 genes implicated in the homozygous phenotype of a proband pedigree to identify candidate mutations, validated them in the family of the proband, studied the functions of the mutant protein, and followed up serum lipid levels after treatment. We discovered that exon 9 c.1268 T>C and exon 8 c.1129 T>G compound heterozygous mutations in the LDLR gene in the proband derived from the mother and father, respectively, in which the mutation of c.1129 T>G has not been reported previously. The mutant LDL-R protein had 57% and 52% binding and internalization functions, respectively, compared with that of the wild type. After 6 months of therapy, the LDL-C level of the proband decreased by more than 50% and the LDL-C of the other family members with heterozygous mutation also reduced to normal. Targeted exome sequencing is an effective method for screening mutation genes in familial hypercholesterolemia. The exon 8 and 9 mutations of the LDLR gene were pedigree mutations. The functions of the mutant LDL-R protein were decreased significantly compared with that of the wild type. Simvastatin plus ezetimibe was proven safe and effective in this preschool-age child.

Hyperpolarization-activated currents (Ih) mediated by hyperpolarization-activated cyclic nucleotide-gated (HCN) channels modulate excitability of myelinated A- and Ah-type visceral ganglion neurons (VGN). Whether alterations in Ih underlie the previously reported reduction of excitability of myelinated Ah-type VGNs following ovariectomy (OVX) has remained unclear. Here we used the intact nodose ganglion preparation in conjunction with electrophysiological approaches to examine the role of Ih remodeling in altering Ah-type neuron excitability following ovariectomy in adult rats. Ah-type neurons were identified based on their afferent conduction velocity. Ah-type neurons in nodose ganglia from non-OVX rats exhibited a voltage 'sag' as well as 'rebound' action potentials immediately following hyperpolarizing current injections, which both were suppressed by the Ih blocker ZD7288. Repetitive spike activity induced afterhyperpolarizations lasting several hundreds of milliseconds (termed post-excitatory membrane hyperpolarizations, PEMHs), which were significantly reduced by ZD7288, suggesting that they resulted from transient deactivation of Ih during the preceding spike trains. Ovariectomy reduced whole-cell Ih density, caused a hyperpolarizing shift of the voltage-dependence of Ih activation, and slowed Ih activation. OVX-induced Ih remodeling was accompanied by a flattening of the stimulus frequency/response curve and loss of PEMHs. Also, HCN1 mRNA levels were reduced by ∼30% in nodose ganglia from OVX rats compared with their non-OVX counterparts. Acute exposure of nodose ganglia to 17beta-estradiol partly restored Ih density and accelerated Ih activation in Ah-type cells. In conclusion, Ih plays a significant role in modulating the excitability of myelinated Ah-type VGNs in adult female rats.

Frontotemporal lobar degeneration (FTLD) represents a group of clinically, neuropathologically and genetically heterogeneous disorders with plenty of overlaps between the neurodegenerative mechanism and the clinical phenotype. FTLD is pathologically characterized by the frontal and temporal lobar atrophy. Frontotemporal dementia (FTD) clinically presents with abnormalities of behavior and personality and language impairments variants. The clinical spectrum of FTD encompasses distinct canonical syndromes: behavioural variant of FTD (bvFTD) and primary progressive aphasia. The later includes nonfluent/agrammatic variant PPA (nfvPPA or PNFA), semantic variant PPA (svPPA or SD) and logopenic variant PPA (lvPPA). In addition, there is also overlap of FTD with motor neuron disease (FTD-MND or FTD-ALS), as well as the parkinsonian syndromes, progressive supranuclear palsy (PSP) and corticobasal syndrome (CBS). The FTLD spectrum disorders are based upon the predominant neuropathological proteins (containing inclusions of hyperphosphorylated tau or ubiquitin protein, e.g transactive response (TAR) DNA-binding protein 43 kDa (TDP-43) and fusedin-sarcoma protein in neurons and glial cells) into three main categories: (1) microtubule-associated protein tau (FTLD-Tau); (2) TAR DNA-binding protein-43 (FTLD-TDP); and (3) fused in sarcoma protein (FTLD-FUS). There are five main genes mutations leading clinical and pathological variants in FTLD that identified by molecular genetic studies, which are chromosome 9 open reading frame 72 (C9ORF72) gene, granulin (GRN) gene, microtubule associated protein tau gene (MAPT), the gene encoding valosin-containing protein (VCP) and the charged multivesicular body protein 2B (CHMP2B). In this review, recent advances on the different clinic variants, neuroimaging, genetics, pathological subtypes and clinicopathological associations of FTD will be discussed.

Background Homozygous familial hypercholesterolaemia is a rare life-threatening disease characterised by markedly elevated LDL cholesterol concentrations and accelerated atherosclerosis. Few studies have focused specifically on this group of patients, especially in China. We set out to investigate the phenotypical characteristics and the genotype–phenotype correlations of homozygous familial hypercholesterolaemia in China. Methods In this analysis of cross-sectional data, we collected data from patients with suspected homozygous familial hypercholesterolaemia from ten clinical hospitals in mainland China from 2003 to 2015. Inclusion criteria included total cholesterol greater than 10 mmol/L, presence of xanthomas, and a family history of premature coronary heart disease. Clinical data and DNA sequence of LDLR, APOB , and PCSK9 were obtained in all patients. All participants gave written informed consent using consent forms approved by the research ethics committee of Beijing Anzhen Hospital, China. Findings We collected data from 64 probands with suspected unrelated homozygous familial hypercholesterolaemia (mean age 13 years [range 2–37]). Molecular diagnosis showed that 15 had homozygous familial hypercholesterolaemia, 34 had compound heterozygous disease, nine had heterozygous disease, and six were mutation-negative. 66 LDLR mutations were found, and no APOB or PCSK9 mutations were found. Of the LDLR mutations, 26 were novel, and the most common mutation, occurring in 11 (17%) of 64 patients, was Trp462X (c1448G→A). The mean total cholesterol concentration was 16·6 mmol/L (SD 3·84; range 10·4–25·9), and the mean LDL cholesterol concentration was 13·86 mmol/L (SD 3·50; range 7·1–23·5). Total cholesterol and LDL cholesterol concentrations did not significantly differ between patients with homozygous familial hypercholesterolaemia, compound heterozygous familial hypercholesterolaemia, heterozygous familial hypercholesterolaemia, or those who were mutation-negative. Interpretation This research provides important clinical and genetic information about suspected homozygous familial hypercholesterolaemia in mainland China. Our data suggest that diagnostic criteria need to be developed to screen patients with potential homozygous familial hypercholesterolaemia. Funding National Natural Science Foundation of China (number 81471098).

To investigate the effect of missense mutation in PCSK9 gene R306S on the metabolism of LDL and to analyze its relationship between structure and function. METHODS The wild type PCSK9 gene and the second or the third grade structures of the mutant R306S coding protein were all forecasted with the aid of Expert Protein Analysis System. The PCSK9 gene eukaryocyte expression vector was constructed and the nucleotide sequence was determined; The pathotype recombinant eukaryotic expression plasmid which carrying PCSK9 gene was constructed by directed-mutation method; Liposorne was used as intermediate to transfect the recombinant plasmid into BEL-7402 cell, the cells receiving no plasmid transfection served as control during this process; The LDL-R level was detected by Western Blot. RESULTS Through simulating the second and the third grade structures of protein, conformations changes were found in the two major coding protein parts of mutated PCSK9 gene: carboxy-terminal domain and catalyzing subunit site domain, the space between these two domains increased as well. And that was not found in wild type of PCSK9 gene. Tansfecting wild type of PCSK9 plasmid into BEL-7402 cell could lead to the decrease of LDL-R mature protein expression compared with controlled group(P

Clostridium butyrium CB2 isolated from chickens was tested as a potential fish probiotic in the Chinese drum, Miichthys miiuy. Fish were fed live (CB), dead CB2 (D-CB) cells (10(8) cells g(-1)) or spent culture supernatant (SCS), for 30 days and challenged with Vibrio anguillarum or Aeromonas hydrophila. Survival was higher in both the CB and the CB-D fed groups, but the SCS group was not significantly different from the control. After feeding live or dead CB2 cells, there was increase in phagocytic activity of the head kidney macrophages, the lysozyme activity of serum and gut mucosa and immunoglobulin (Ig) level. The SCS group showed no obvious change in immune parameters. The results suggest that live or dead Clostridium butyrium CB2 has an immunomodulatory effect on fish.

We investigated the effects of fish protein hydrolysate (FPH) on growth performance and humoral immune response of the large yellow croaker (Pseudosciaena crocea R.). One thousand and two hundred large yellow croakers [initial average weight: (162.75+/-23.85) g] were divided into four groups and reared in floating sea cages (3 m x 3 m x 3 m). The animals were fed with 4 diets: basal diet only (control) or diets supplemented with 5%, 10% and 15% (w/w) FPH. The results show that dietary FPH levels significantly influenced the growth and immunity of the large yellow croaker. Compared with the control group, total weight gain (TWG) in all treatment groups, relative weight gain (RWG) and specific growth rate (SGR) in fish fed with diets supplemented with 10% and 15% FPH were significantly increased (P0.05). Similar results were observed in immune parameters [lysozyme activity, serum complements, immunoglobulin M (IgM)]. Lysozyme activity, complement C4 and IgM were also significantly increased (P0.05) in fish fed with diets supplemented with 10% and 15% FPH, while complement C3 level was significantly increased (P0.05) in all treatment groups. In general, with the supplementation of FPH, particularly at dose of 10%, the growth performance and immunity of the large yellow croaker can be improved effectively.