Objective To study the prevalence of the mtDNA A1555G gene mutation in Chinese population with nonsyndromic hearing impairment.Methods PCR-RFLP,directional sequencing of PCR products were applied in 325 patients with nonsyndromic hearing impairment and 50 normal controls.Results The mutation rate of the mtDNA A1555G was 14.5% (47/325),28 of 47 cases were homozygosis,19 of 47 cases were heterozygosis.The same mutation was not detected in the control subjects.Conclusion The mutation rate of the mtDNA A1555G is relatively high in the Chinese NSHI patients,the mutation type includes both heterozygosis and homozygosis.

OBJECTIVETo study mitochondrial DNA (mtDNA) A1555G mutation in seven families with nonsyndromic hearing loss (NSHL).

METHODSPolymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and real time-amplification refractory mutation system-quantitative PCR (ARMS-qPCR) were applied to detect mtDNA A1555G mutation in seven NSHL families. Related clinical data were also collected and analyzed.

RESULTSThe mtDNA A1555G mutation was detected in members from the maternal side, including heteroplasmy and homozygosis, others were negative for this mutation. The copy number of homoplasmic or heteroplasmic mutations of mtDNA A1555G correlated well with the degree of deafness (R = 0.341, P = 0.022 and R = 0.85, P = 0.015, respectively).

CONCLUSIONThe mutation rate of the mtDNA A1555G is high in the NSHL patients, the mutation type include heteroplasmy and homozygosis. There is significant correlation between the mtDNA A1555G copy number and the severity of hearing loss.

Adolescent ; Adult ; Aged ; Aged, 80 and over ; Child, Preschool ; DNA, Mitochondrial ; genetics ; Female ; Gene Dosage ; Hearing Loss ; genetics ; pathology ; Humans ; Infant ; Male ; Middle Aged ; Pedigree ; Point Mutation ; Young Adult

This study was aimed to develop a simple, rapid, accurate and wound-free single-tube multiplex polymerase chain reaction (PCR) assay, which can be used for molecular screening and prenatal diagnosis of 3 types of the commonest deletional alpha-thalassemias in Chinese population. The deletional alpha-thalassemias free fetal DNA of 50 anemic pregnant plasma were detected by means of single-tube multiplex PCR with 4 groups of primers designed by using the gap-PCR protocol. The results showed that 5 cases were found to be alpha-thalassemias in free fetal DNA of 50 anaemia pregnant plasma, out of them 3 cases were the Southeast Asia type of deletional alpha-thalassemias, 2 cases were alpha-(3.7) type of deletional alpha-thalassemias (rightward deletion). It is concluded that the single-tube multiplex PCR assay presented in this study is a simple and convenient, rapid and accurate method for detecting 3 common deletional alpha-thalassemias, and use of this method has an important significance for the deletional alpha-thalassemias to ensure the population health quality and reduce the social burden.
DNA ; genetics ; DNA Primers ; Female ; Fetus ; Humans ; Polymerase Chain Reaction ; methods ; Pregnancy ; Prenatal Diagnosis ; Sequence Deletion ; alpha-Thalassemia ; diagnosis ; genetics

BACKGROUNDIt has been suggested that the ratio of mutant and wild type mitochondrial DNA may be related to its clinical phenotype. In this study, we developed a high sensitive real-time amplification refractory mutation system-quantitative polymerase chain reaction (RT-ARMS-qPCR) assay for quantitation of the mitochondrial DNA (mtDNA) with a mutated 1555 site, and explored the relationship between the ratio of mutated mtDNA and the severity of hearing loss of mitochondrial deafness (MD) patients of Fujian province in China.

METHODSAn amplified mtDNA fragment containing the 1555 site was ligated into a vector to construct a plasmid DNA standard. An RT-ARMS-qPCR system was used to measure the mtDNA copy number containing wild-type and mutant sequences in a cohort of 126 MD patients of Fujian province in China. Combined with the clinical data, we explored the relationship between the ratio of mutated mtDNA and the severity of hearing loss of MD.

RESULTSThe variation coefficient in the cohort was 1.21%, the interassay variation coefficient was 1.78%, and the linear range was 10(2) - 10(8) copies/µl for detecting a recombinant, wild-type plasmid. The primers amplified only the intended sequences. Mutation copy number correlated with the degree of deafness in sporadic cases with heteroplasmic mutations of mtDNA A1555G (R = 0.811, P = 0.003), but not in sporadic cases with homoplasmic mutations (R = 0.007, P = 0.989). The copy number of homoplasmic or heteroplasmic mutations of mtDNA A1555G in familial cases correlated with degree of deafness (R = 0.352, P = 0.023 and R = 0.90, P = 0.012, respectively).

CONCLUSIONSThe RT-ARMS-qPCR system is suitable for determining the copy number of mtDNA fragments containing the A1555G mutation. The ratio of mutated mtDNA correlates with the severity of hearing loss of MD.

Adolescent ; Adult ; Child ; Child, Preschool ; China ; DNA, Mitochondrial ; genetics ; Deafness ; genetics ; Female ; Hearing Loss ; genetics ; Humans ; Male ; Mutation ; Reverse Transcriptase Polymerase Chain Reaction ; Young Adult

BACKGROUNDThe capsule associated protein 10 gene (cap10) is indispensible for the formation of the polysaccharide capsule, and is important in maintaining virulence of the Cryptococcus (C.) neoformans. In this study, we aimed to construct an short hairpin RNA (shRNA) expression vector targeting C. neoformans cap10 gene expression and confirm its biologic relevance.

METHODSA pair of oligonucleotides targeting the cap10 cDNA sequence was designed and synthesized. It was cloned into the plasmid psilencer4.1-CMV neo to construct an eukaryotic shRNA expression vector. The vector was transfected into C. neoformans cells using the LiAc method. The expression of cap10 was assessed by real-time fluorescence quantitative PCR. Groups of C. neoformans cells were incubated with murine macrophage-like J774A.1 cells, and the phagocytic indexes and ratios were determined by the microscopic observation method.

RESULTSThe expression of cap10 in C. neoformans cells transfected with ps4.1 neo-cap10 ((175,535.00 ± 47,004.00) copies/µl) was lower than that of cells transfected with the empty vector ((512,698.89 ± 32,318.02) copies/µl) and mock transfected cells ((562,931.66 ± 65,928.41) copies/µl). The average phagocytic ratio and phagocytic index of J774A.1 cells following incubation with C. neoformans were higher for cells transfected with ps4.1 neo-cap10 (0.21 ± 0.02, (19.06 ± 1.66)%) than for the control experimental group (0.08 ± 0.02, (6.57 ± 1.23)%) and the blank experimental group ((0.07 ± 0.01), (5.89 ± 1.07)%) (P < 0.05).

CONCLUSIONSThe cap10 shRNA vector was successfully prepared and transfected into C. neoformans cells. The effect of RNA interference on the expression of the C. neoformans cap10 gene is effective, and it can induce phagocytosis of C. neoformans.

Animals ; Cell Line ; Cryptococcus neoformans ; genetics ; metabolism ; Fungal Proteins ; genetics ; metabolism ; Genetic Vectors ; genetics ; Phagocytosis ; Plasmids ; genetics ; Polymerase Chain Reaction ; RNA Interference ; RNA, Small Interfering ; genetics ; Transfection

BACKGROUNDMutations in the cationic trypsinogen gene (PRSS1) have been detected in patients with hereditary pancreatitis (HP). This study investigated the prevalence of the R122H (c.365 G > A), A121T (c.361 G > A) and D162D (c.488 C > T) mutations or polymorphisms in the common, non-hereditary forms of chronic pancreatitis and in an HP family.

METHODSDNA was prepared from blood samples of 54 patients with chronic pancreatitis (35 alcoholic, 17 idiopathic and 2 hereditary) and 120 normal controls. The PRSS1 genes were amplified by polymerase chain reaction (PCR) and their products were analyzed by sequencing and related clinical data were also collected.

RESULTSA new polymorphism (c.488 C > T) of PRSS1 was found in 25 patients with chronic pancreatitis (including one affected member of the HP family) and six members of the normal controls. The C/T genotype was significantly increased in chronic pancreatitis (OR: 16.379, 95% CI: 5.7522 - 52.3663), the frequency of c.488 C > T change was in according with the Hardy-Weinberg equilibrium, but it doesn't affect the clinical phenotype. The commonly reported change of R122H (c.365 G > A) was not detected in any of the study subjects. c.361 G > A was found in 2 affected members and one unaffected carrier in an HP family. One of the affected members of an HP family had c.361 G > A mutation and polymorphism (c.488 C > T) in the PRSS1 gene at the same time. The patient's clinical values (C3, C4, CA19-9 and HbA1c) were higher than those of the other patients with chronic pancreatitis. The two patients with HP developed diabetes mellitus and their father died with pancreatic cancer.

CONCLUSIONA new polymorphism (c.488 C > T) in the PRSS1 gene is associated with chronic pancreatitis, but it did not affect the clinical phenotype while the A121T (c.361 G > A) mutation in the gene shows a significant correlation in the patients with HP.

Female ; Humans ; Male ; Mutation ; Pancreatitis ; genetics ; Pancreatitis, Chronic ; genetics ; Polymorphism, Genetic ; Trypsin ; Trypsinogen ; genetics

BACKGROUNDA high mortality rate of pancreatic cancer becomes a bottleneck for further treatment with long-term efficacy. It is urgent to find a new mean to predict the early onset of pancreatic cancer accurately. The authors hypothesized that genetic variants of cationic trypsinogen (PRSS1) gene could affect trypsin expression/function and result in abnormal activation of protease activated receptor-2 (PAR-2), then lead to pancreatic cancer. The aim of this study was to elaborate some novel mutations of PRSS1 gene in the patients with pancreatic cancer.

METHODSTotally 156 patients with pancreatic cancer and 220 unrelated individuals as controls were enrolled in this study. The mutations of PRSS1 gene were analyzed by direct sequencing. K-ras Mutation Detection Kit was used to find the general k-ras gene disorder in the pancreatic cancer tissue. Then the clinical data were collected and analyzed simultaneously.

RESULTSThere were two patients who carried novel mutations which was IVS 3 + 157 G > C of PRSS1 gene in peripheral blood specimens and pancreatic cancer tissue. What's more, it was surprising to find a novel complicated mutation of exon 3 in PRSS1 gene (c.409 A > G and c.416 C > T) in another young patient. The complicated mutation made No. 135 and No. 137 amino acid transfer from Thr to Ala and Thr to Met respectively. No any mutation was found in the normal controls while no mutations of k-ras gene were detected in the three patients.

CONCLUSIONMutations of PRSS1 gene may be an important factor of pancreatic cancer.

Adult ; Asian Continental Ancestry Group ; Female ; Humans ; Male ; Mutation ; Pancreatic Neoplasms ; genetics ; Trypsin ; genetics

OBJECTIVESTo evaluate the efficacy of intrameatal application of low dosage alprostadil (PGE1) cream (300 mcg) for the treatment of erectile dysfunction (ED).

METHODSA total of 43 ED patients were selected in the study based on the inclusion criteria. All of the patients signed informed consent forms and entered a 4-week open-label clinical study. A dosage of 300 mg PGE1 in 75 mg cream was applied intrameatally.

RESULTSThe results showed that the primary efficacy (IIEF Q3 + Q4) reached 70.73% after application of the cream. The successful intercourse rate was 86.41%. Based on the GAQ (global assessment Question); 73.17% of the patients were satisfied with their sexual life. At the same time, all of the secondary criteria supported the primary efficacy results. Two patients withdrew during the study period. Six patients (14.63%) had urethral pain or penile redness, which were mostly mild and transient.

CONCLUSIONSWith intrameatal low dosage (300 mcg PGE1) of the PGE1 cream can achieve an equivalent efficacy as that with the full dosage.

Adult ; Aged ; Alprostadil ; administration & dosage ; therapeutic use ; Erectile Dysfunction ; drug therapy ; Humans ; Male ; Middle Aged ; Penile Erection ; drug effects ; Treatment Outcome ; Vasodilator Agents ; administration & dosage ; therapeutic use

Golgi protein 73 (GP73) is a transmembrane protein on the Golgi apparatus and can be cut and released into the blood. In recent years, an increasing number of clinical studies have shown that the elevated serum GP73 level is closely related to liver diseases. And thus GP73 is expected to be used as a new serum marker for assessing progress of chronic liver diseases. Herein, the clinical application of serum GP73 in chronic hepatitis, liver fibrosis, liver cirrhosis and hepatocellular carcinoma with different etiologies was reviewed based on available literatures; and a research outlook in this field is made.
Biomarkers ; Carcinoma, Hepatocellular ; Golgi Apparatus ; Humans ; Liver Cirrhosis ; Liver Neoplasms