To investigate the effect of GFP fused to C terminal of integrin alpha(IIb) on the biosynthesis and expression of alpha(IIb) beta(3) compound, the alpha(IIb) GFP expression plamid, named palpha(IIb) GFP, the cDNA of alpha(IIb) was constructed from p3.1-2b and fused to pEGFP-N1 in frame. When the sequence of palpha(IIb) GFP was confirmed by sequencing it was transferred to Chinese Hamster Ovary (CHO) cells with or without p3.1-3a expressing integrin beta(3). Then the expression of alpha(IIb) GFP fusion protein was confirmed by Western blot and then its subcellular localization was determined with laser confocal scanning microscopy. The results showed that the target gene was cloned into recombinant vector by restriction analysis and sequencing. Overexpression of the fusion protein in the transfected CHO cells was identified with Western blot. Subcellular localization analysis confirmed that alpha(IIb) GFP was expressed in CHO cells and could be transferred from endoplasmic reticulum to Golgi apparatus. It is concluded that the eukaryotic expression plasmid containing alpha(IIb) GFP fusion gene is successfully constructed. GFP fused to the cytoplasmic tail of integrin alpha(IIb) allows the normal expression of alpha(IIb) beta(3) in CHO cells.
Animals ; Blotting, Western ; CHO Cells ; Cricetinae ; Cricetulus ; Endoplasmic Reticulum ; metabolism ; Golgi Apparatus ; metabolism ; Green Fluorescent Proteins ; genetics ; metabolism ; Microscopy, Confocal ; Platelet Glycoprotein GPIIb-IIIa Complex ; genetics ; metabolism ; Recombinant Fusion Proteins ; genetics ; metabolism ; Transfection

Abstract: To analyze the clinical, therapeutic and laboratory characteristics of disseminated cryptococcosis caused by Cryptococcus neoformans invading the blood stream in patient with liver cirrhosis and splenectomy. A 30-year-old male underwent splenectomy plus pericardial devascularization due to "splenomegaly and hypersplenism" in March in 2016. The patient had intermittent fever after operation for many times, and successively accompanied with back pain, left lower limb abscess and right hip pain. The highest body temperature was 39 ℃. CT and MRI revealed the lung lesion and multiple bone destruction. During that period, the effect of antibiotics was not good. On April 19th, 2017, Gram's stain, India ink stain, API 32C, Vitek 2 Compact, ribosomal ITS and IGS sequence analysis were performed to identify the strain isolated from the pus and blood stream. The serum of the patient was detected for cryptococcal antigen. Antifungal susceptibility test was used to determine drug sensitivity and minimum inhibitory concentration (MIC). The Cryptococcus neoformans isolated from fresh pus specimen showed a prominent, thick capsule after India ink stain. The colonies isolated from pus and blood stream were identified Cryptococcus neoformans using API 32C, Vitek 2 Compact, and sequence analysis of rDNA ITS and IGS. Cryptococcal capsule antigen was positive. The minimal inhibitory concentrations of 5-Flucytosine, amphotericin B, fluconazole, itriconazole, voriconazole against the isolate were <4 μg/mL, <0.5 μg/mL, 4 μg/mL, ≤0.25 μg/mL, 0.125 μg/mL respectively. The patient was initially treated with intravenous amphotericin B and flucytosine. After anti-Cryptococcus treatment for two months, the patient clinically improved, and the lesions were reduced on a follow-up CT scan. The patient made a full functional recovery after treatment for six months. Cryptococcosis has hidden onset, atypical clinical symptoms and lack of specificity. Blood stream is the main channel for Cryptococcus to spread and involve many organs of the whole body, including skin, bone and so on. Therefore, early use of blood culture to monitor blood flow dissemination, actively removing the primary focus and cutting off the infection route in time and carrying out effective anti-Cryptococcus treatment are conducive to the patient's early recovery.

OBJECTIVETo study the effect of glycoprotein (GP) alpha II bA2334C mutation on the biosynthesis and expression of alpha II bbeta3 complex.

METHODSThe GP alpha II bA2334C eukaryotic expression plasmid pc3.1-2334M2b was constructed. Chinese hamster ovary (CHO) cells were transfected with the plasmid with or without integrin beta3 expression plasmid pc3.1-3a. The whole expression of alpha II bA2334C was confirmed by Western blot and the membrane expression was analyzed by flow cytometry. A newly constructed alpha II bA2334C GFP fusion protein expressing plasmid was used to determine its subcellular localization by laser confocal scanning microscopy.

RESULTSExpression of the mutant protein, alpha II bA2334C, in the transfected CHO cells was confirmed by Western blot with a lower rate of the mature type than the wild type control. The expression on membrane was only 25% of the normal. Subcellular localization analysis showed that alpha II bA2334C GFP was able to be expressed in CHO cells and could be transported from endoplasmic reticulum to Golgi apparatus.

CONCLUSIONSThe mutant alpha II bA2334C can be synthesized in CHO cells and form alpha II bbeta3 complex. However, only a small fraction of the premature alpha II bA2334C can be transported to Golgi apparatus and transformed to mature alpha II b. The possible pathogenesis of this type II thrombasthenia may be that the misfolded alpha II bA2334C is partially degraded in the endoplasmic reticulum causing lower expression of alpha II bbeta3 complex on the membrane and resulting in impared function of platelets than normal alpha II b.

Animals ; Biological Transport ; Blotting, Western ; CHO Cells ; Cricetinae ; Cricetulus ; Female ; Green Fluorescent Proteins ; genetics ; metabolism ; Humans ; Liposomes ; Microscopy, Confocal ; Middle Aged ; Mutation ; Plasmids ; genetics ; Platelet Glycoprotein GPIIb-IIIa Complex ; genetics ; metabolism ; Transfection

OBJECTIVECarnitine deficiency has been associated with progressive cardiomyopathy due to compromised energy metabolism. The objective of this study was to investigate clinical features of carnitine deficiency-induced cardiomyopathy and the therapeutic efficacy of L-carnitine administration.

METHODBetween January 2010 and December 2011, filter-paper blood spots were collected from 75 children with cardiomyopathy. Free carnitine and acylcarnitine profiles were measured for each individual by tandem mass spectrometry (MS/MS). For those in whom carnitine deficiency was demonstrated, treatment was begun with L-carnitine at a dose of 150 - 250 mg/(kg·d). Clinical evaluation, including physical examination, electrocardiography, chest x-ray, echocardiography and tandem mass spectrometry, was performed before therapy and during follow-up.

RESULTOf 75 cardiomyopathy patients, the diagnosis of carnitine deficiency was confirmed in 6 patients, which included 1 boy and 5 girls. Their age ranged from 0.75 to 6 years. Free carnitine content was (1.55 ± 0.61) µmol/L (reference range 10 - 60 µmol/L). Left ventricular end-diastolic diameter (LVDd) was (5.04 ± 0.66) cm and left ventricular ejection fraction (LVEF) was (38.5 ± 10.5)%. After 10 - 30 d therapy of L-carnitine, free carnitine content rose to (30.59 ± 15.02) µmol/L (t = 4.79, P < 0.01). LVDd decreased to (4.42 ± 0.67) cm (t = 4.28, P < 0.01) and LVEF increased to (49.1 ± 7.6)% (t = 6.59, P < 0.01). All patients received follow-up evaluations beyond 6 months of treatment. Clinical improvement was dramatic. LVEF returned to normal completely in all the 6 patients. LVDd decreased further in all the 6 patients and returned to normal levels in 3 patients. No clinical signs or symptoms were present in any of the 6 patients. The only complications of therapy had been intermittent diarrhea in 1 patient.

CONCLUSIONTandem mass spectrometry is helpful to diagnose carnitine deficiency and should be performed in all children with cardiomyopathy. L-carnitine has a good therapeutic effect on carnitine deficiency-induced cardiomyopathy.

Adolescent ; Cardiomyopathies ; diagnosis ; drug therapy ; etiology ; Cardiotonic Agents ; administration & dosage ; therapeutic use ; Carnitine ; blood ; deficiency ; therapeutic use ; Child ; Child, Preschool ; Electrocardiography ; Female ; Follow-Up Studies ; Humans ; Infant ; Male ; Retrospective Studies ; Tandem Mass Spectrometry ; Treatment Outcome ; Ventricular Function, Left ; drug effects

OBJECTIVEVerverck index (VI) reflects thoracic development, body type, and nutritional status. This study aimed to investigate the VI of singleton neonates with a gestational age (GA) of 27-42 weeks at birth, and to establish percentile curves of VI of the neonates.

METHODSCross-sectional cluster sampling was performed between April 2013 and September 2015. Body weight, body length, and chest circumference were measured for 16 865 singleton neonates with a GA of 27-42 weeks in two hospitals in Shenzhen, China. VI was calculated and the percentile curves of VI were plotted for the neonates.

RESULTSMean VIs were obtained for singleton neonates with a gestational age of 27-42 weeks (in three groups of male, female, and both sexes), and related 3rd-97th percentile curves were plotted. As for the 50th percentile curve, the singleton neonates with a GA of 27 weeks had the lowest 50th percentile value of VI, which gradually increased with the increase in GA. The singleton neonates with a GA of 42 weeks had the highest 50th percentile value of VI. Girls had a slightly higher 50th percentile value of VI than boys in all GA groups.

CONCLUSIONSVI of neonates increases with the increase in GA. Female neonates may have a slightly better thoracic development, body type, and nutritional status than male neonates at birth. The percentile curves of VI plotted for singleton neonates with a GA of 27-42 weeks (in three groups of male, female, and both sexes) can provide a basis for evaluating thoracic development, body type, and nutritional status of neonates at birth in Shenzhen, China.

OBJECTIVETo establish the intrauterine growth curves of neonates in Shenzhen, China and to investigate the intrauterine growth of neonates in Shenzhen.

METHODSCross-sectional cluster sampling was performed for an on-the-spot investigation of 16 887 neonates (9 418 males and 7 469 females) with a gestational age of 27-42 weeks who were born in two hospitals in Shenzhen from April 2013 to September 2015. The Lambda Mu Sigma (LMS) method was used for the curve fitting of body weight, body length, head circumference, chest circumference, and crown-rump length.

RESULTSThe 3rd-97th percentile intrauterine growth curves for body weight, body length, head circumference, chest circumference, and crown-rump length were plotted for the neonates with a gestational age of 27-42 weeks who were divided into three groups (male, female, and mixed). The male neonates had significantly higher curves for the five indices than the female counterparts. The pattern and changing trend of body weight curves of these neonates were basically consistent with those in China Neonatal Network.

CONCLUSIONSThe percentile intrauterine growth curves for body weight, body length, head circumference, chest circumference, and crown-rump length in neonates with a gestational age of 27-42 weeks in Shenzhen which has been established can provide a reference for clinical practice in the department of neonatology.

Body Height ; Body Weight ; Cross-Sectional Studies ; Crown-Rump Length ; Female ; Fetal Development ; physiology ; Gestational Age ; Head ; anatomy & histology ; Humans ; Infant, Newborn ; Male ; Thorax ; anatomy & histology

OBJECTIVETo explore the effects of obesity on the peak level of luteinizing hormone (LH) in the gonadotropin-releasing hormone (GnRH) agonist test and obesity-related hormones in girls with central precocious puberty (CPP).

METHODSThree hundred and thirty-three girls with CPP who underwent the GnRH agonist test between 2012 and 2014 were classified into three groups: normal weight (n=123), overweight (n=108), and obesity (n=102), according to body mass index (BMI). The sexual development indices were compared between the three groups. Twenty girls were randomly selected from each group for evaluation of the serum levels of leptin, sex hormone binding globulin (SHBG), neurokinin B, and kisspeptin. The correlation of BMI with the levels of various hormones was assessed using Pearson correlation analysis.

RESULTSThere was no significant difference in mean age at diagnosis between the three groups; however, the bone age was significantly higher in the overweight and obesity groups than in the normal weight group (P<0.05). The peak level of LH in the GnRH agonist test and SHBG level in the normal weight group were significantly higher than those in the overweight and the obesity groups, while the serum levels of leptin and neurokinin B were significantly lower in the normal weight group than in the overweight and the obesity groups (P<0.05). BMI was negatively correlated with the peak level of LH in the GnRH agonist test and SHBG level (P<0.05), and positively correlated with the levels of leptin and neurokinin B (P<0.05).

CONCLUSIONSThe effects of BMI on the result of the GnRH agonist test and levels of obesity-related hormones should be taken into account in girls with precocious puberty.

Body Mass Index ; Child ; Female ; Gonadotropin-Releasing Hormone ; agonists ; Humans ; Leptin ; blood ; Luteinizing Hormone ; blood ; Neurokinin B ; blood ; Obesity ; blood ; Puberty, Precocious ; blood ; Sex Hormone-Binding Globulin ; analysis

OBJECTIVETo investigate the situation of normal blood pressure and prevalence and risk factors of prehypertension in Han, Uygur and Hazakh population ≥ 35 years old of Xinjiang Uygur autonomous region.

METHODSRandom samples were used to analyze the incidence of normal blood pressure and prehypertension in 3 nationalities based on data from Han, Uygur and Hazakh residents living in 7 areas (Urumqi, Ke lamayi, Fukang, the Turfan Basin locality, Hetian locality, Altay locality, and Yili Hazakh autonomous prefecture) in Xinjiang Uygur autonomous region.

RESULTSA total of 16 460 subjects were selected with cluster sampling, and eligible data of 14 618 subjects were actually analyzed in Han, Uygur and Hazakh population ≥ 35 years old of Xinjiang Uygur autonomous region. Incidence of normal blood pressure was 24.9% and the prevalence of prehypertension was 34.24% (35.6% fro male and 31.5% for female, P < 0.01) in Han, Uygur and Hazakh population ≥ 35 years old in Xinjiang Uygur autonomous region. The prevalence of prehypertension was 37.34%, 32.95% and 30.62% in Han, Uygur and Hazakh population, respectively. The prevalence of prehypertension decreased with increasing age in Han and Uygur population ≥ 45 years old, and in Hazakh population ≥ 35 years old (all P < 0.05). Multiple logistic regression analysis revealed that hypertriglyceridemia ≥ 1.7 mmol/L, age ≥ 45 years old, fasting plasma glucose ≥ 7.0 mmol/L and body mass index ≥ 24.0 kg/m(2) were risk factors while the female was a protective factor of prehypertension.

CONCLUSIONThe people with normal blood pressure is few and the prevalence of prehypertension is high in Han, Uygur and Hazakh population ≥ 35 years old of Xinjiang Uygur autonomous region, and it is associated with many risk factors.

Adult ; Aged ; Asian Continental Ancestry Group ; Blood Pressure ; China ; epidemiology ; Cross-Sectional Studies ; Female ; Humans ; Incidence ; Male ; Middle Aged ; Prehypertension ; epidemiology ; physiopathology ; Prevalence ; Risk Factors ; Surveys and Questionnaires

OBJECTIVETo study the Kaup index (KI), an index used to evaluate body burliness and nutritional status, of neonates with a gestational age (GA) of 27-42 weeks at birth, and to establish the percentile curves of KI.

METHODSCross-sectional cluster sampling was performed from April 2013 to September 2015 to select 16 887 singleton neonates with a GA of 27-42 weeks in two hospitals in Shenzhen, China. Body weight and body length were measured to calculate KI. The percentile curves of KI were plotted in these neonates.

RESULTSMean KIs and corresponding standard deviations were obtained for singleton neonates with a gestational age of 27-42 weeks (in male, female, and mixed groups), and the 3rd-97th percentile curves of KI were plotted. The singleton neonates with a GA of 27 weeks had the lowest 50th percentile value of KI, and KI gradually increased with GA. Boys had a higher 50th percentile value of KI than girls in each GA group. In all groups except the 33-week GA group, boys had a higher mean KI than girls, and there was a significant difference in the mean KI between boys and girls in the GA groups of 34 and 36-40 weeks (P<0.05).

CONCLUSIONSKI of neonates at birth increases with GA, suggesting that body density and body burliness increase with GA. Boys have better body burliness than girls at birth. The percentile curves of KI plotted for singleton neonates with a GA of 27-42 weeks (in male, female, and mixed groups) can provide a reference for evaluating the body burliness and nutritional status of neonates at birth in Shenzhen.

Cross-Sectional Studies ; Female ; Gestational Age ; Humans ; Infant, Newborn ; Male ; Nutritional Status

OBJECTIVETo establish the intrauterine growth curve of twin neonates, and to investigate the intrauterine growth status of twin neonates.

METHODSCross-sectional cluster sampling was performed for an on-the-spot investigation of 1 296 live twin neonates who were born in two hospitals in Shenzhen between April 2013 and September 2015. The Lambda-Mu-Sigma method was used for the curve fitting of body weight, body length, head circumference, chest circumference, and crown-rump length.

RESULTSThe means and 3rd-97th percentile intrauterine growth curves for body weight, body length, head circumference, chest circumference, and crown-rump length were obtained for the 1 296 twin neonates with a gestational age of 27-40 weeks. The curve values of the 1 296 twin neonates for body weight, body length, head circumference, chest circumference, and crown-rump length were all lower than those of singleton neonates in Shenzhen that had been reported, and the difference increased with increasing gestational age.

CONCLUSIONSThe intrauterine growth curves for body weight, body length, head circumference, chest circumference, and crown-rump length of twin neonates with a gestational age of 27-40 weeks in Shenzhen obtained in this study can provide a reference for evaluating the intrauterine growth status of twin neonates among the current population in Shenzhen.

Body Height ; Body Weight ; Cross-Sectional Studies ; Female ; Fetal Development ; Gestational Age ; Humans ; Infant, Newborn ; Male ; Pregnancy ; Twins