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The purpose of this study is to apply Fuzzy Synthesis Judge to set up a model of performance evaluation criterion used to assess the quality of enterprise's outsourcing management. This study adopts means of literature review and expert-based interviews to contribute to an adequate evaluation criteria used to measure the performance of outsourcing activities. In terms of data collection and analysis, the participants consist of experts in aviation industry. By means of questionnaire distribution to experts, the data analysis is applied with fuzzy synthesis judge to examine the weight value. Consequently, this study utilizes fuzzy synthesis judge to qualify the performance evaluation and determine the optimal model used to examine the efficiency of outsourcing management. This study offers a model of evaluation criterion which makes it possible for enterprises to make the best outsourcing performance.

Objective: To perform molecular diagnosis and pedigree analysis for one case with α-thalassemia who does not conform to the genetic laws, and explore the effects of a newly discovered rare mutation ( HBA2:c.*12G>A ) on clinical phenotypes., Methods: Blood samples of the proband and her family members were collected for blood routine analysis, and the hemoglobin components were analyzed by capillary electrophoresis. The common α- and β-globin gene loci in Chinese population were detected by conventional techniques (Gap-PCR, RDB-PCR). The α-globin gene sequences ( HBA1, HBA2 ) were analyzed by Sanger sequencing., Results: By analyzing the test results of proband and her family members, the genotype of the proband was -α 3.7 / HBA2:c.*12G>A , her father was HBA2:c.*12G>A heterozygous mutation carrier., Conclusion: This study identifies a rare α-globin gene mutation ( HBA2:c.*12G>A ) that has not been reported before. It is found that heterozygous mutation carriers present with static α-thalassemia.

Objective: To investigate two cases of rare pathogenic genes, initiation codon mutations in HBA2 gene, combined with Southeast Asian deletion and their family members to understand the relationship of HBA2:c.2T>C and HBA2:c.2delT mutations with clinical phenotype., Methods: The peripheral blood of family members was obtained for blood cell analysis and capillary electrophoresis hemoglobin analysis. Gap-PCR and reverse dot blotting (RDB) were used to detect common types of mutations in ɑ-thalassaemia gene. Sanger sequencing was used to analyze HBA1 and HBA2 gene sequence., Results: Two proband genotypes were identified as --SEA/αα with HBA2:c.2T>C and --SEA/αα with HBA2:c.2delT . HBA2:c.2T>C/WT and HBA2:c.2delT/WT was detected in family members. They all presented with microcytic hypochromic anemia., Conclusion: When HBA2:c.2T>C and HBA2:c.2delT are heterozygous that can lead to static α-thalassemia phenotype, and when combined with mild α-thalassemia, they can lead to the clinical manifestations of hemoglobin H disease. This study provides a basis for genetic counseling.

Purpose: In this study, a large diameter visible trephine was designed and used in percutaneous endoscopic lumbar interbody fusion to increase endoscopic bone decompression efficiency. Large diameter visible trephine-related technical notes and preliminary clinical experience are described., Methods: A large diameter visible trephine was designed with normal diameter visible trephine as template. A total of 38 patients with lumbar degenerative diseases who underwent single-level percutaneous endoscopic lumbar interbody fusion with large or normal diameter visible trephine were included into a retrospective study. Operation time, bone decompression time, blood loss, intraoperative fluoroscopy, bone decompression fluoroscopy, and dura or nerve injury cases were recorded and analyzed statistically. Visual Analog Scale (VAS) scores and Oswestry Disability Index (ODI) were used to analyze the clinical outcomes of the 2 groups., Results: The baseline data of the 2 groups were statistically similar. There was no significant difference in postoperative VAS and ODI scores between the 2 groups. Operation time and bone decompression time of large diameter visible trephine group were significantly shorter than that of normal diameter visible trephine group (P < 0.05). Intraoperative fluoroscopy times and bone decompression fluoroscopy times of large diameter visible trephine group were significantly more than that of normal diameter visible trephine group (P < 0.05). Blood loss of the 2 groups were not statistically different. There were no dura or nerve injury cases in the 2 groups., Conclusions: For percutaneous endoscopic lumbar interbody fusion, the large diameter visible trephine is a safe and efficient endoscopic bone decompression tool under fluoroscopic guidance., (Copyright © 2022 Elsevier Inc. All rights reserved.)

An understanding of the epidemiology of poststroke dementia (PSD) is necessary to inform research, practice and policy. With increasing primary studies, a contemporary review of PSD could allow for analyses of incidence and prevalence trends. Databases were searched using a prespecified search strategy. Eligible studies described an ischaemic or mixed stroke cohort with prospective clinical assessment for dementia. Pooled prevalence of dementia was calculated using random-effects models at any time after stroke (primary outcome) and at 1 year (range: 6-18 months), stratified for inclusion of prestroke dementia. Meta-regression explored the effect of year of study. Sensitivity analyses removed low-quality or outlier studies. Of 12 505 titles assessed, 44 studies were included in the quantitative analyses. At any time point after stroke, the prevalence of PSD was 16.5% (95% CI 10.4% to 25.1%) excluding prestroke dementia and 22.3% (95% CI 18.8% to 26.2%) including prestroke dementia. At 1 year, the prevalence of PSD was 18.4% (95% CI 7.4% to 38.7%) and 20.4% (95% CI 14.2% to 28.2%) with prestroke dementia included. In studies including prestroke dementia there was a negative association between dementia prevalence and year of study (slope coefficient=-0.05 (SD: 0.01), p<0.0001). Estimates were robust to sensitivity analyses. Dementia is common following stroke. At any point following stroke, more than one in five people will have dementia, although a proportion of this dementia predates the stroke. Declining prevalence of prestroke dementia may explain apparent reduction in PSD over time. Risk of dementia following stroke remains substantial and front-loaded, with high prevalence at 1 year post event., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2022. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Objective: To evaluate the clinical value of aspirin as a prophylactic for transplant renal artery stenosis (TRAS). Methods: From January 2017 to November 2019, clinical data of 307 patients who had undergone renal transplant in Zhengzhou University People's Hospital were collected. Patients were divided into two groups: the treatment group (124 recipients who had taken oral aspirin 100 mg/d after transplant) and the control group (183 recipients who had not taken aspirin after transplant). The general data, incidence of initially diagnosed and confirmed TRAS, type of renal artery anastomosis vessels, duration of stenosis, location of stenosis, and complications were compared between the two groups. The treatment group was further divided into two subgroups, the early group (92 recipients) and the delayed group (32 recipients), according to the time of starting aspirin after operation. Subgroup analysis was performed. Results: Among all 307 patients included, there were 241 males and 66 females, aged 19-64 years. There were no statistical difference between the treatment and control groups in terms of gender, age, comorbidities, number of arterial vessels, type of graft, and acute rejection all P >0.05. Among 46 initially diagnosed TRAS patients, 13 (10.5%) and 33 (18.0%) cases were in the treatment and control group respectively, with no statistically significant difference in stenosis rate ( P >0.05). The number of confirmed TRAS patients was 1 (0.8%) and 24 (13.1%) in the treatment and control group respectively, with statistically significant difference in stenosis rate ( P< 0.001). The proportion of patients with bleeding disorders in the treatment group was slightly higher than that in the control group (13.7% vs 8.7%), and the proportion of infarct diseases was slightly lower than that in the control group (1.6% vs 4.9%). But there was no significant difference in aspirin-related complications between the two groups ( P >0.05). Subgroup analysis showed that there was no significant difference in initially diagnosed and confirmed TRAS and aspirin-related complications between the early group and the delayed group (all P >0.05). Conclusions: Oral low-dose aspirin after kidney transplantation can effectively reduce the incidence of TRAS, without increasing the risk of aspirin-related complications.

Objective: To proceed the clinical evaluation of DNA microarray for thalassemia gene detection., Methods: Peripheral blood samples of 166 thalassemia gene test subjects were collected and tested for thalassemia genes by microarray chip method and Gap-PCR method combined with PCR-reverse dot blot hybridization method according to double-blind control test. The specificity, sensitivity, positive predictive value, negative predictive value, and total coincidence rate of the microarray chip method were evaluated. When the two methods were inconsistent, multiplex ligation dependent probe amplification (MLPA) was used to verify the deletional α-thalassemia., Results: Compared with Gap-PCR method, specificity, sensitivity, positive predictive value, negative predictive value, Youden index, and total coincidence rate of microarray chip method was 100% (70/70), 96.88% (93/96), 100% (93/93), 95.89% (70/73), 0.969, and 97.59% (162/166), respectively, while compared with PCR-reverse dot blot hybridization method was 100% (125/125), 100% (41/41), 100% (41/41), 100% (125/125), 1, and 100% (166/166), respectively., Conclusion: The microarray chip method for α-thalassemia gene detection shows the advantages of high specificity, sensitivity, and throughput.

Objective: To perform dried blood spots thalassemia gene detection in patients with positive blood phenotypes by microarray technology, and evaluate its value in clinical detection., Methods: DNA samples were extracted from dried blood spots of 410 patients. Microarray technology was used to detect 3 deletion and 3 non-deletion types of α-thalassemia and 19 β-thalassemia point mutations which were common gene mutions in China., Results: There were 357 positive cases in all the 410 tested samples with the positive rate 87.07%, among which 299 cases (72.93%) carried deletion or point mutations of α-thalassemia, 29 cases (7.07%) carried point mutations of β-thalassemia and 29 cases (7.07%) carried gene mutations of complex αβ-thalassemia syndrome. The mutations of α-thalassemia were involved with -- SEA heterozygous deletion (177 cases, 59.2%), α CS heterozygote (60 cases, 20.07%) and several other genotypes. The common mutations of β- thalassemia were involved with β CD41-42 heterozygote (10 cases, 34.48%) and β CD17 heterozygote (9 cases, 31.03%). The mutations of complex αβ-thalassemia syndrome were mainly involved with -- SEA /αα+β CD17 /β N (7 cases, 24.14%), α CS α/αα + β CD41-42 /β N (3 cases, 10.34%) and -α 4.2 /αα + β CD17 /β N (3 cases, 10.34%)., Conclusion: The most common genetic mutations are -- SEA for α-thalassemia and CD41-42 for β-thalassemia in Liuzhou, Guangxi Zhuang Autonomous Region. A and β-thalassemia can be detected at the same time by microarray chip technology in a high throughput manner.

Cadmium (Cd) contamination of rice is a serious food safety issue that has recently been gaining significant public attention. Therefore, reduction of Cd accumulation in rice grains is an important objective of rice breeding. The use of favourable alleles of Cd accumulating genes using marker-assisted selection (MAS) is theoretically feasible. In this study, we validated a segment covering OsHMA3-OsNramp5-OsNramp1 on chromosome 7 of japonica for establishing low-cadmium accumulating indica rice variety. The OsHMA3-OsNramp5-OsNramp1 jap haplotype significantly decreased grain Cd concentration in middle-season indica genetic background. The improved 9311 carrying the OsHMA3-OsNramp5-OsNramp1 jap haplotype with recurrent parent genome recovery of up to 91.6% resulted in approximately 31.8% decrease in Cd accumulation in the grain and with no penalty on yield. There is a genetic linkage-drag between OsHMA3-OsNramp5-OsNramp1 jap and the gene conditioning heading to days (HTD) in the early-season indica genetic background. Because the OsHMA3-OsNramp5-OsNramp1-Ghd7 jap haplotype significantly increases grain Cd concentration and prolongs growth duration, the linkage-drag between OsHMA3-OsNramp5-OsNramp1 and Ghd7 should be broken down by large segregating populations or gene editing. A novel allele of OsHMA3 was identified from a wide-compatibility japonica cultivar, the expression differences of OsNramp1 and OsNramp5 in roots might contribute the Cd accumulating variation between japonica and indica variety.

The aim of this study was to investigate a family with nonhomologous sequence recombination of HBA1 and HBA2 genes and provide a favorable basis for genetic counseling and eugenics. Peripheral blood of family members was collected. Hematological parameters were determined by an automated cell counter and hemoglobin (Hb) analysis was performed using high performance liquid chromatography (HPLC). Villus samples were taken for prenatal diagnosis (PND). Gap-polymerase chain reaction (gap-PCR) and reverse dot-blot were used for thalassemia genotyping. DNA sequencing was used to analyze the gene sequence of HBA1 (α1-globin) and HBA2 (α2-globin). The nonhomologous sequence recombination allelic variant of HBA1 and HBA2 genes were identified, namely, a gene conversion on the HBA2 gene called α12 ( HBA12 ). The α12 allele consists primarily of the HBA2 gene sequence except for a segment of the IVS-II in which HBA2 -specific sequences have been replaced by HBA1 -specific sequences. The following genotypes were observed: - - SEA /αα12 (Southeast Asian deletion), αα/αα12 and α QS α/αα12 (Hb Quong Sze or Hb QS; HBA2 : c.377T>C), and all manifested as small cell hypochromic anemia. To find the α12 allele in the Chinese population and clarify the influence of the α12 allele and its common inheritance with abnormal Hb and α-thalassemia (α-thal) on α-globin gene expression can help guide clinical diagnosis and genetic counseling.

Objective: To investigate the clinical effects of percuteneous endoscopic interlaminar discectomy(PEID)and fenestration discectomy(FD) for the treatment of L₅S₁ lumbar disc herniation(LDH)., Methods: A retrospective analysis was made on 86 patients with L₅S₁ LDH from January 2014 to March 2017 and followed up. According to the different surgical methods, the patients were divided into PEID group(43 cases) and FD group(43 cases). All operations were performed under general anesthesia. Forty-three patients in PEID group underwent percuteneous endoscopic interlaminar discectomy (PEID) and other 43 patients in FD group underwent classical fenestration discectomy (FD). Operative incision, operative time, intraoperative blood loss, duration of hospitalization, time of lying in bed after surgery, complication were compared between two groups. Pre- and postoperative visual analogue scale(VAS) of affected extremity pain and lumbar pain were recorded. Postoperative creatine kinase (CK) was observed in two groups. Modified Macnab criteria was used to evaluate the clinical effects. MRI was used to observe the survival rate of paraspinal muscle after operation., Results: The length of skin incision, intraoperative blood loss, duration of hospitalization, time of lying in bed after surgery of PEID group and FD group were(0.7±0.1) cm, (8.0±3.0) ml, (3.0±1.5) d, (1.0±0.5) d and(5.0±1.8) cm, (62.0±50.5) ml, (11.0±2.5) d, (3.0±0.8) d, there was significant differences between two groups( P <0.05). VAS of affected extremity at 24 hours and 1 year after operation was obviously decreased in two groups( P <0.05), but there was no significant difference between groups( P >0.05). VAS of lumbar pain in PEID group and FD group were respectively (2.99±0.32), (5.44±1.31) scores at 24 hours after operation, and (1.56±0.60), ( 3.05±0.24) at 1 year after operation, there was significant differences between two groups( P <0.05). CK at 24, 48 hours after operation of FD group were obviously increased( P <0.05). According the modified Macnab criteria to evaluate the clinical effect, the rate of excellent and good of PEID group and FD group were 93% and 95%, respectively. The survival rate of paraspinal muscle by MRI in PEID group at 1 year after operation was higher than that in FD group( P <0.05). No complications such as spinal dura mater tearing, nerve root injury, vascular injury, intervertebral space infection were found in two groups., Conclusions: Both of the two methods are safe and can obtain satisfactory effect, but PEID is more in line with concept of minimally invasive and has more advantages in paraspinal muscle protection, operative incision, intraoperative blood loss, duration of hospitalization, time of lying in bed after operation., Competing Interests: The authors of this article and the planning committee members and staff have no relevant financial relationships with commercial interests to disclose., (Copyright© 2019 by the China Journal of Orthopaedics and Traumatology Press.)

Objective: This study aims to elucidate the potential role of microRNA-448 in the recovery of spinal cord injury (SCI), and to explore the underlying mechanism., Materials and Methods: MicroRNA-448 expression was determined by microarray and the established SCI model in mice. The target gene of microRNA-448 was predicted using bioinformatics. The functional binding of the target gene to microRNA-448 was verified by Dual-Luciferase reporter gene assay. The regulatory effects of microRNA-448 and Bcl-2 on apoptosis, motor neuron number and grip strength were evaluated. After injection of microRNA-448 mimics, microRNA-448 inhibitor or Bcl-2 siRNA in mice, expression levels of PI3K/AKT and Caspase3 were detected by quantitative Real Time-Polymerase Chain Reaction (qRT-PCR) and Western blot., Results: Grip strength of SCI mice significantly decreased compared with mice in the sham group. The microRNA-448 expression gradually increased with the progression of SCI, whereas the Bcl-2 expression decreased. Dual-Luciferase reporter gene assay showed the binding condition between microRNA-448 and Bcl-2. Furthermore, the Bcl-2 expression was negatively regulated by microRNA-448 at both mRNA and protein levels. The injection of microRNA-448 inhibitor into the injured spinal cord of SCI mice significantly upregulated the expressions of p-PI3K, p-AKT and Caspase3, as well as motor neuron regeneration and grip strength. However, the promotive effects of microRNA-448 inhibitor were blocked by Bcl-2 siRNA transfection., Conclusions: MicroRNA-448 is upregulated after SCI, which may be involved in the regenerative process of spinal motor nerves by regulating PI3K/AKT/Bcl-2 axis.

In this study, we aimed to reveal the role of miR-191 in apoptosis of renal tubular epithelial cells and in the involvement of renal ischemia-reperfusion injury. Renal transplantation rat model was established. miR-191 and Cystathionine-β-synthase (CBS) were measured by qRT-PCR and Western blot. The regulation of miR-191 on CBS was detected by luciferase reporter assay. We found miR-191 expression in platelets and platelet microvesicles (P-MVs) of patients and model rats was significantly upregulated than that of health and normal rats. Also, mRNA and protein levels of CBS in renal tissues of patients were significantly downregulated than that of health and normal rats. We also found that P-MVs could transfer miR-191 to HK-2 cells. Luciferase reporter assay showed that CBS was a direct target of miR-191. In addition, we proved that P-MVs-secreted miR-191 inhibited CBS expression in HK-2 cells, and P-MVs-secreted miR-191 promoted HK-2 cell apoptosis via CBS. Finally, we verified the trends of CBS expressions, HK-2 cell apoptosis and apoptosis-related proteins in vivo were similar as the trends in vitro. Therefore, CBS was a direct target of miR-191, and miR-191 could transfer to HK-2 cells via P-MVs to decrease the expression of CBS, thus to promote cell apoptosis and renal IR injury.

This study aimed to analyze the clinical phenotype of chromosome 9p deletion or duplication and its relationship with karyotype. A patient, female, aged 6 months, visited the hospital due to motor developmental delay. Karyotype analysis identified abnormalities of chromosome 9 short arm, and high-throughput sequencing found 9p24.3-9p23 deletion and 9p23-9p13.1 duplication. Her parents had a normal karyotype. Karyotype analysis combined with high-throughput sequencing is of great significance for improving the efficiency of etiological diagnosis in children with motor developmental delay or multiple congenital deformities and mental retardation.

Objective: Microvesicles (MVs) are plasmalemmal vesicles that are released from various cells and regarded as a mediator of intermolecular communication. In present study, we aimed to evaluate the therapeutic efficacy of the bone marrow mesenchymal stem cells (BM-MSCs)-derived MVs in the mice kidney transplant model and explored the underlying mechanism., Methods: BM-MSCs were isolated from C57BL/6 mice and identified using flow cytometry. In vivo allogenic kidney transplantation model of mice was performed between C57BL/6 mice (recipient) and BALB/c mice (donor). Recipient-type BM-MSC (0.1ml) or equal volume of medium as a control was injected i.v. 24h after kidney transplantation. Serum was collected for creatinine concentration detection at 14 d after transplantation. Dendritic cells (DCs) phenotype and miR-146a expression level in plant was identified. Immature DCs (iDCs) and mature DCs (mDCs) were derived from monocytes. MVs were separated from BM-MSCs., Results: BM-MSCs positive for CD29 (95.8%) and CD44 (94.7%) were cultured and confirmed to prolong the allogenic kidney graft survival in mice. Importantly, the expression of miR-146a increased significantly in DCs of BM-MSCs-treated allogenic kidney. Moreover, both BM-MSCs and MVs derived from BM-MSCs enhanced miR-146a expression in iDCs and mDCs in vitro. Furthermore, MVs substantially reduced IL-12 mRNA expression and IL-12 production of mDCs whereas this action was reversed by miR-146a silencing. MiR-146a silencing also abrogated the MVs-induced decrease in serum creatinine, reduction of immature DCs phenotype in transplant and increase in miR-146a expression level., Conclusion: In summary, our data suggested that the BM-MSCs-derived MVs improved allogenic kidney transplantation survival through inhibiting DCs maturity by miR-146a., (Copyright © 2017 European Federation of Immunological Societies. Published by Elsevier B.V. All rights reserved.)

Objective: To study the gene mutation profile of primary carnitine deficiency (PCD) in neonates, and to provide a theoretical basis for early diagnosis and treatment, genetic counseling, and prenatal diagnosis of PCD., Methods: Acylcarnitine profile analysis was performed by tandem mass spectrometry using 34 167 dry blood spots on filter paper. The SLC22A5 gene was sequenced and analyzed in neonates with free carnitine (C0) levels lower than 10 μmol/L as well as their parents., Results: In the acylcarnitine profile analysis, a C0 level lower than 10 μmol/L was found in 10 neonates, but C0 level was not reduced in their mothers. The 10 neonates had 10 types of mutations at 20 different sites in the SLC22A5 gene, which included 4 previously unreported mutations: c.976C>T, c.919delG, c.517delC, and c.338G>A. Bioinformatics analysis showed that the four new mutations were associated with a risk of high pathogenicity., Conclusions: Tandem mass spectrometry combined with SLC22A5 gene sequencing may be useful for the early diagnosis of PCD. Identification of new mutations enriches the SLC22A5 gene mutation profile.

Background: Spinal muscular atrophy (SMA) is mainly caused by deletions in SMA-related genes. The objective of this study was to develop gene-dosage assays for diagnosing SMA., Methods: A multiplex, quantitative PCR assay and a CNVplex assay were developed for determining the copy number of SMN1, SMN2, and NAIP. Reproducibility and specificity of the two assays were compared to a multiple ligation-dependent probe amplification (MLPA) assay. To evaluate reproducibility, 30 samples were analyzed three times using the three assays. A total of 317 samples were used to assess the specificity of the two assays., Results: The multiplex quantitative PCR (qPCR) assay had higher reproducibility. Intra-assay CVs were 3.01%-8.52% and inter-assay CVs were 4.12%-6.24%. The CNVplex assay had ratios that were closer to expected (0.49-0.5 for one copy, 1.03-1.0 for two copies, and 1.50-1.50 for three copies). Diagnostic accuracy rates for the two assays were 100%., Conclusions: The multiplex qPCR assay was a simple, rapid, and cost-effective method for routine SMA diagnosis and carrier screening. The CNVplex assay could be used to detect SMAs with complicated gene structures. The assays were reliable and could be used as alternative methods for clinical diagnosis of SMA.

The aim of the study was to analyze the treatment effect of transumbilical single-port laparoscopic varicocelectomy (TUSPLV) on recurrent varicocele (VC). In order to compare the surgical effects of TUSPLV to traditional retroperitoneal ligation of the internal spermatic vein, 64 patients with recurrent VC were enrolled and divided into the control group (n=30) and the observation group (n=34). Patients in the control group underwent surgery using traditional retroperitoneal ligation of the internal spermatic vein, while those in the observation group underwent surgery using TUSPLV. The results showed that the time of operation and bleeding volume in the observation group were significantly lower. The occurrence and recurrence rates of periprocedural complications were considerably lower in the observation group. Differences were statistically significant (P<0.05). In terms of the pregnancy rate, the difference between the 2 groups had no statistical significance (P>0.05). We concluded that employing TUSPLV to treat recurrent VC was safe and effective.

This study aimed to identify the type of carnitine palmitoyltransferase 2 (CPT2) gene mutation in the child with carnitine palmitoyltransferase II (CPT II) deficiency and her parents and to provide the genetic counseling and prenatal diagnosis for the family members. As the proband, a 3-month-old female baby was admitted to the hospital due to fever which had lasted for 8 hours. Tandem mass spectrometric analysis for blood showed an elevated plasma level of acylcarnitine, which suggested CPT II deficiency. The genomic DNA was extracted from peripheral blood of the patient and her parents. Five exon coding regions and some intron regions at the exon/intron boundaries of the CPT2 gene were analyzed by PCR and Sanger sequencing. Amniotic fluid was taken from the mother during the second trimester, and DNA was extracted to analyze the type of CPT2 gene mutation. Sanger sequencing results showed that two mutations were identified in the CPT2 gene of the proband: c.886C>T (p.R296X) and c.1148T>A (p.F383Y), which were inherited from the parents; the second child of the mother inherited the mutation of c.886C>T (p.R296X) and showed normal acylcarnitine spectrum and normal development after birth. It is concluded that the analysis of CPT2 gene mutations in the family suggested that the proband died of CPT II deficiency and that the identification of the mutations was helpful in prenatal diagnosis in the second pregnancy.

Some studies have indicated that the Wnt/β-catenin signaling pathway is activated following spinal cord injury, and expression levels of specific proteins, including low-density lipoprotein receptor related protein-6 phosphorylation, β-catenin, and glycogen synthase kinase-3β, are significantly altered. We hypothesized that methylprednisolone treatment contributes to functional recovery after spinal cord injury by inhibiting apoptosis and activating the Wnt/β-catenin signaling pathway. In the current study, 30 mg/kg methylprednisolone was injected into rats with spinal cord injury immediately post-injury and at 1 and 2 days post-injury. Basso, Beattie, and Bresnahan scores showed that methylprednisolone treatment significantly promoted locomotor functional recovery between 2 and 6 weeks post-injury. The number of surviving motor neurons increased, whereas the lesion size significantly decreased following methylprednisolone treatment at 7 days post-injury. Additionally, caspase-3, caspase-9, and Bax protein expression levels and the number of apoptotic cells were reduced at 3 and 7 days post-injury, while Bcl-2 levels at 7 days post-injury were higher in methylprednisolone-treated rats compared with saline-treated rats. At 3 and 7 days post-injury, methylprednisolone up-regulated expression and activation of the Wnt/β-catenin signaling pathway, including low-density lipoprotein receptor related protein-6 phosphorylation, β-catenin, and glycogen synthase kinase-3β phosphorylation. These results indicate that methylprednisolone-induced neuroprotection may correlate with activation of the Wnt/β-catenin signaling pathway., Competing Interests: Conflicts of Interest: None declared.

X-linked ichthyosis (XLI) is a metabolic disease with steroid sulfatase deficiency and often occurs at birth or shortly after birth. The encoding gene of steroid sulfatase, STS, is located on the short arm of the X chromosome, and STS deletion or mutation can lead to the development of this disease. This study collected the data on the clinical phenotype from a family, and the proband, a boy aged 11 years with full-term vaginal delivery, had dry and rough skin and black-brown scaly patches, mainly in the abdomen and extensor aspect of extremities. Peripheral blood samples were collected from each family member and DNA was extracted. Multiplex ligation-dependent probe amplification (MLPA) was used to measure the copy number of STS on the X chromosome. Whole-genome microarray was used to determine the size of the segment with microdeletion in the X chromosome. MLPA was then used for prenatal diagnosis for the mother of the proband. The results revealed that the proband and another two male patients had hemizygotes in STS deletion. Gene microarray identified a rare deletion with a size of 1.6 Mb at Xp22.31 (chrX: 6,516,735-8,131,442). Two female family members were found to be carriers. Prenatal diagnosis showed that the fetus carried by the proband's mother was a carrier of this microdeletion. This study showed STS gene deletion in this family of XLI, which causes the unique skin lesions of XLI. MLPA is a convenient and reliable technique for the molecular and prenatal diagnosis of XLI.

A 9-day-old male patient was admitted to the hospital because of cough, anhelation, feeding difficulty and lethargy. The diagnostic examinations indicated pulmonary infection, severe metabolic acidosis, hyperglycemia, hyperammonemia and pancytopenia in the patient. Blood and urine screening and isovaleryl-CoA dehydrogenase (IVD) gene detection for inherited metabolic diseases were performed to clarify the etiology. Tandem mass spectrometric screening for blood showed an elevated isovalerylcarnitine (C5) level. The organic acid analysis of urine by gas chromatography-mass spectrometry showed significantly increased levels in isovaleryl glycine and 3-hydroxyisovaleric acid. Homozygous mutations (c.1208A>G, p.Tyr403Cys) in the IVD gene were identified in the patient. His parents were heterozygous carriers. After the treatment with low-leucine diets and L-carnitine for 3 days, the patient showed a significant improvement in symptoms, but he died one week later. It is concluded that the neonates with pneumonia and metabolic decompensation of unknown etiology should be screened for genetic metabolic disease.

Medium- and short-chain acyl-CoA dehydrogenase deficiency is a disorder of fatty acid β-oxidation. Gene mutation prevents medium- and short-chain fatty acids from entry into mitochondria for oxidation, which leads to multiple organ dysfunction. In this study, serum acylcarnitines and the organic acid profile in urea were analyzed in two children whose clinical symptoms were hypoglycemia and metabolic acidosis. Moreover, gene mutations in the two children and their parents were evaluated. One of the patients was a 3-day-old male who was admitted to the hospital due to neonatal asphyxia, sucking weakness, and sleepiness. The serum acylcarnitine profile showed increases in medium-chain acylcarnitines (C6-C10), particularly in C8, which showed a concentration of 3.52 μmol/L (reference value: 0.02-0.2 μmol/L). The analysis of organic acids in urea gave a normal result. Sanger sequencing revealed a reported c.580A>G (p.Asn194Asp) homozygous mutation at exon 7 of the ACADM gene. The other patient was a 3-month-old female who was admitted to the hospital due to cough and recurrent fever for around 10 days. The serum acylcarnitine profile showed an increase in serum C4 level, which was 1.66 μmol/L (reference value: 0.06-0.6 μmol/L). The analysis of organic acids in urea showed an increase in the level of ethyl malonic acid, which was 55.9 (reference value: 0-6.2). Sanger sequencing revealed a reported c.625G>A (p.Gly209Ser) homozygous mutation in the ACADS gene. This study indicates that screening tests for genetic metabolic diseases are recommended for children who have unexplained metabolic acidosis and hypoglycemia. Genetic analyses of the ACADM and ACADS genes are helpful for the diagnosis of medium- and short-chain acyl-CoA dehydrogenase deficiency.

Objective: We compared a three-drug combination therapy (control group) consisting of tacrolimus, mycophenolate mofetil, and corticosteroids in living donor renal transplantation with a four-drug combination therapy (study group), in which the doses of tacrolimus and mycophenolate mofetil were halved and the immunosuppressive drug mizoribine was added, in order to determine whether the incidence rates of acute rejection after transplantation between the study group and the control group are similar, whether the study group regimen prevents the occurrence of calcineurin inhibitor-induced renal damage, and whether the study group regimen prevents adverse effects such as diarrhea caused by mycophenolate mofetil., Methods: We investigated the incidence of acute rejection, serum creatinine levels, and estimated glomerular filtration rate and the incidence of adverse effects such as diarrhea., Results: There was no significant difference between the two groups in the incidence of acute rejection. Renal function (estimated glomerular filtration rate and serum creatinine) was maintained in the control group whereas in the study group renal function gradually improved, with a statistical difference observed at 12 months. The incidence of gastrointestinal symptoms including diarrhea was significantly higher in the control group than in the study group. There was no significant difference in the incidence of cytomegalovirus infection and other adverse effects., Conclusion: These results suggest the study group therapy is an effective regimen in preventing acute rejection and the deterioration of renal function. These results also show this therapy can reduce the incidence of adverse effects such as gastrointestinal symptoms.

Background: Spinal muscular atrophy (SMA) is caused by SMN1 dysfunction, and the copy number of SMN2 and NAIP can modify the phenotype of SMA. The aim of this study was to analyze the copy numbers and gene structures of SMA-related genes in Chinese SMA patients and unrelated healthy controls., Methods: Forty-two Chinese SMA patients and two hundred and twelve unrelated healthy Chinese individuals were enrolled in our study. The copy numbers and gene structures of SMA-related genes were measured by MLPA assay., Results: We identified a homozygous deletion of SMN1 in exons 7 and 8 in 37 of 42 patients (88.1%); the other 5 SMA patients (11.9%) had a single copy of SMN1 exon 8. The proportions of the 212 unrelated healthy controls with different copy numbers for the normal SMN1 gene were 1 copy in 4 individuals (1.9%), 2 copies in 203 (95.7%) and 3 copies in 5 (2.4%). Three hybrid SMN genes and five genes that lack partial sequences were found in SMA patients and healthy controls. Distributions of copy numbers for normal SMN2 and NAIP were significantly different (P < 0.001) in people with and without SMA., Conclusion: The copy numbers and gene structures of SMA-related genes were different in Chinese SMA patients and healthy controls.

Background: The Southeast Asian deletion (--(SEA)) is the most commonly observed mutation among diverse α-thalassemia alleles in Southeast Asia and South China. It is generally argued that mutation --(SEA), like other variants causing hemoglobin disorders, is associated with protection against malaria that is endemic in these regions. However, little evidence has been provided to support this claim., Results: We first examined the genetic imprint of recent positive selection on the --(SEA) allele and flanking sequences in the human α-globin cluster, covering a genomic region spanning ~410 kb, by genotyping 28 SNPs in a Chinese population consisting of 76 --(SEA) heterozygotes and 138 normal individuals. The pattern of linkage disequilibrium (LD) and the long-range haplotype test revealed a signature of positive selection. The network of inferred haplotypes suggested a single origin of the --(SEA) allele., Conclusions: Thus, our data support the hypothesis that the --(SEA) allele has been subjected to recent balancing selection, triggered by malaria.

Prostate cancer is a leading cause of death in male populations across the globe. With the advent of gene expression arrays, many microarray studies have been conducted in prostate cancer, but the results have varied across different studies. To better understand the genetic and biologic mechanisms of prostate cancer, we conducted a meta-analysis of two studies on prostate cancer. Eight key genes were identified to be differentially expressed with progression. After gene co-expression analysis based on data from the GEO database, we obtained a co- expressed gene list which included 725 genes. Gene Ontology analysis revealed that these genes are involved in actin filament-based processes, locomotion and cell morphogenesis. Further analysis of the gene list should provide important clues for developing new prognostic markers and therapeutic targets.

Reliable detection of large deletions from cell-free fetal DNA (cffDNA) in maternal plasma is challenging, especially when both parents have the same deletion owing to a lack of specific markers for fetal genotyping. In order to evaluate the efficacy of a non-invasive prenatal diagnosis (NIPD) test to exclude α-thalassemia major that uses SNPs linked to the normal paternal α-globin allele, we established a novel protocol to reliably detect paternal SNPs within the (--(SEA)) breakpoints and performed evaluation of the diagnostic potential of the protocol in a total of 67 pregnancies, in whom plasma samples were collected prior to invasive obstetrics procedures in southern China. A group of nine SNPs identified within the deletion breakpoints were scanned to select the informative SNPs in each of the 67 couples DNA by multiplex PCR based mini-sequencing technique. The paternally inherited SNP allele from cffDNA was detected by allele specific real-time PCR. A protocol for reliable detection of paternal SNPs within the (--(SEA)) breakpoints was established and evaluation of the diagnostic potential of the protocol was performed in a total of 67 pregnancies. In 97% of the couples one or more different SNPs within the deletion breakpoint occurred between paternal and maternal alleles. Homozygosity for the (--(SEA)) deletion was accurately excluded in 33 out of 67 (49.3%, 95% CI, 25.4-78.6%) pregnancies through the implementation of the protocol. Protocol was completely concordant with the traditional reference methods, except for two cases that exhibited uncertain results due to sample hemolysis. This method could be used as a routine NIPD test to exclude gross fetal deletions in α-thalassemia major, and could further be employed to test for other diseases due to gene deletion.

Screening for carriers of spinal muscular atrophy (SMA) is necessary for effective clinical/prenatal diagnosis and genetic counseling. However, a population-based study of SMA prevalence in mainland China has not yet been conducted. In this study, the copy number of survival motor neuron (SMN) genes was determined in 1712 newborn cord blood samples collected from southern China and from 25 core families, which included 26 SMA patients and 44 parents, to identify SMA carriers. The results presented 13 groups with different SMN1/SMN2 ratios among 1712 newborn individuals, which corresponded to 1535 subjects with two copies of SMN1, 119 with three copies of SMN1, 17 with four copies of SMN1, and 41 with a heterozygous deletion of SMN1 exon 7. Simultaneously, two '2+0' genotypes and two point mutations were found among the 44 obligate carriers in the core families, including a novel SMN1 splice-site mutation that was identified in the junction between intron 6 and exon 7 (c. 835-1G>A). These results indicated that the carrier frequency is 1/42 in the general Chinese population and that duplicated SMN1 alleles and de novo deletion mutations are present in a small number of SMA carriers. In addition, we developed and validated a new alternative screening method using a reverse dot blot assay for rapid genotyping of deletional SMA. Our research elucidated the genetic load and SMN gene variants that are present in the Chinese population, and could serve as the basis for a nationwide program of genetic counseling and clinical/prenatal diagnosis to prevent SMA in China.

Glucose-6-phosphate dehydrogenase (G6PD) deficiency is a common inherited disorder worldwide including southern China. We developed and validated a reverse dot blot (RDB) assay for the rapid and simultaneous genotyping of six mutations (c.95A>G, c.871G>A, c.1004C>T, c.1024C>T, c.1376G>T and c.1388G>A), that were common mutations in the Chinese G6PD deficiency population, and one polymorphism (c.1311C>T). Reliable genotyping of wild-type and mutant genomic DNA samples was achieved by means of a test strip onto which allele-specific oligonucleotide probe lines are fixed in parallel. This method involves a multiplex PCR amplification of three fragments in the G6PD target sequence and a manual hybridization/detection protocol. The entire procedure starting from blood sampling to the identification of mutations requires less than 6 h. The diagnostic reliability of this reverse dot blot assay was evaluated on 207 pre-typed samples by using direct DNA sequence analysis in a blind study. The reverse dot blot typing was in complete concordance with the reference method. The reverse dot blot assay was proved to be a simple, rapid, highly accurate, and cost-effective method to identify common G6PD mutations in Chinese population.

Objective: To study the role of the glucocorticoid receptor (GR) signal pathway and downstream cytokine interleukin-6 (IL-6) in androgen-independent growth of prostate cancer (PC)., Methods: The human androgen-dependent PC (ADPC) cell line LNCaP and androgen-independent PC (AIPC) cell line DU145 were cultured in vitro. Immunocytochemistry was used to examine the expression of the androgen receptor (AR), GR, HSP90 and IL-6. The GR antagonist RU486 was used to treat cultured cells, and the effects of RU486 on the proliferation of both cell lines were analyzed by MTT assay. Expression of HSP90 and IL-6 mRNA and protein was assessed by RT-PCR and Western blots, respectively., Results: LNCaP cells were AR-positive and GR-negative, whereas DU145 cells were GR-positive and AR-negative. The expression of HSP90 and IL-6 in DU145 cells were significantly stronger than that in LNCaP cells (p < 0.01). RU486 had no obvious effects on the growth of LNCaP cells, but exerted a significant time- and dose-dependent growth inhibition on DU145 cells at doses as low as 0.1 micromol/l. RU486 treatment of DU145 cells also resulted in a dose-dependent decrease in the expression of HSP90 and IL-6 mRNA and protein., Conclusions: The GR signal pathway may be the main survival pathway for DU145 cells. Abnormal hyperactivation of the GR signal pathway and its promoting the expression of HSP90 and IL-6 contribute to the progression of ADPC to AIPC after androgen ablation., ((c) 2008 S. Karger AG, Basel.)

There have been conflicting results obtained when comparing the in vitro with in vivo effects of prostaglandin E (PGE) on immune function. The in vitro studies have demonstrated immune suppression with PGE administration while the in vivo studies demonstrated improved survival when utilizing infected models. To attempt to resolve this discrepancy, we evaluated the in vivo effect of PGE on host immune function utilizing multiple rat models. PGE was found to have no effect on the ability of leucocytes to infiltrate a sponge matrix wound over a 2-week period of study. PGE also failed to alter the percentage of T-lymphocyte subset populations infiltrating the sponge matrix model. There was noted to be no effect of PGE on the ability of neutrophils to chemiluminescence, or on metabolic function of lymphocytes. In conclusion, PGE does not appear to have immunosuppressive properties when studied using certain in vivo models.

Patients suffering severe trauma frequently become immunosuppressed following injury. This can predispose patients to infectious sequelae. Biochemically, these patients synthesize excessive quantities of cyclooxygenase products (prostaglandins). It has been hypothesized that the prostaglandins cause the immunosuppression and that inhibition of the cyclooxygenase enzyme could thus prevent the immunosuppression. We investigated the effect of the cyclooxygenase inhibitor ibuprofen on the inflammatory response. Rats were subjected to a 30% total body surface area burn and were administered either ibuprofen for a period of 7 days or 14 days, or were administered the carrier for 14 days. The rats were then killed and multiple immunologic variables were measured. Ibuprofen was found to decrease neutrophil chemiluminescence, lymphocyte blastogenesis, and helper/inducer T-lymphocyte infiltration of a sponge matrix model. The same ibuprofen protocol decreased survival in a cecal ligation and puncture model. In conclusion, the cyclooxygenase enzyme system appears to produce metabolites essential for optimal survival following traumatic injury.

Background: Insulinlike growth factor 1 (IGF-1) has previously been demonstrated to improve the nutritional status of burned animals. The method by which it achieves this result has not yet been fully elucidated, but may be the result of alterations in hepatic metabolism., Objective: To determine if IGF-1 is able to correct the burn-induced impairments in hepatic metabolic function., Design: Seventy-two Sprague-Dawley rats were subjected to a sham burn (n = 24), or a 50% total body surface area scald burn (n = 48). Half the scald burn group received 3 micrograms/kg per day of IGF-1. The remainder received a placebo. The rats were sequentially assayed for multiple components of hepatic function., Results: Insulinlike growth factor 1 corrected the burn-induced decrease in hepatic adenosine triphosphate concentration and prevented the burn-induced increase in hepatic ketone body levels. Insulinlike growth factor 1 was also able to prevent the burn-induced decrease in the hepatic acetoacetate-beta-hydroxybutyrate ratio. Since this ratio is directly proportional to mitochondrial redox potential this indicates that IGF-1 is also able to prevent the burn-induced impairment in hepatic redox potential., Conclusions: These data indicate that part of the previously demonstrated beneficial effect of IGF-1 in burn injury may be due to its ability to improve multiple components of hepatic metabolism.

Oncology patients suffer multiple detrimental metabolic alterations. Among these are catabolism of tumor free body mass to supply nutrients to feed the tumor. This results not only in enhanced tumor growth but also poor wound healing and immunosuppression of the tumor host. Efforts are therefore being directed at finding methods for improving the nutritional status of the tumor host without enhancing tumor growth. We investigated the ability of two hormones, insulin-like growth factor-1 (IGF-1) and insulin, to improve physiologic function in tumor-bearing animals. Tumor-bearing animals received a continuous infusion of IGF-1 (2.20 mg/kg/day), insulin (820 microns/kg/day) or placebo via an osmotic minipump for 7 days. All animals were pair fed to eliminate nutritional intake as a variable. The placebo group lost 31.37 +/- 4.3 g of tumor free body mass during the study period. The insulin treated group lost 26.34 +/- 7.42 g and the IGF-1 group lost 5.07 +/- 3.25 g (P < 0.001, ANOVA). IGF-1 treatment failed to alter plasma glucose, lactate, or total amino acid concentration and failed to alter hepatic ketone body concentrations, but did improve hepatic mitochondria redox potential. Finally, IGF-1 improved splenic weight by 110% and splenic lymphocyte count by 300%. In conclusion IGF-1 appears to offer potential in supporting tumor free host body mass without stimulating tumor growth.

Cyclooxygenase products are believed to be a major regulator of host tumor necrosis factor-alpha (TNF-alpha) production in response to trauma and sepsis. To study this relationship, Lewis rats underwent a 30% burn or sham burn. Dimethyl-prostaglandin E (dPGE, 50 micrograms/kg), ibuprofen (IFU, 2 mg/kg), or saline was administered twice daily. Rats were sacrificed at Day 7 to obtain Kupffer cells, peritoneal macrophages, splenic macrophages, and neutrophils. For in vivo studies, 10(6) cells from each group were cultured with 10 micrograms of lipopolysaccharide (LPS). For in vitro studies, cells from the burn and sham groups were cultured with LPS and dPGE (10 micrograms/ml), IBU (10 micrograms/ml), or saline. The supernatants were harvested after 2, 6, and 24 hr of culture and assayed for TNF-alpha (mu/ml) by L929 cytolysis. Burn injury resulted in a significant increase in Kupffer cell and neutrophil TNF-alpha production compared to the sham group (P < 0.001, ANOVA). The administration of IBU to burned animals led to a pronounced elevation of TNF-alpha production by Kupffer cells, peritoneal macrophages, and neutrophils compared to vehicle-treated burned animals (P < 0.001, ANOVA). With in vitro studies, IBU increased Kupffer cell, peritoneal macrophage, and neutrophil TNF-alpha release by 213, 327, and 198%, respectively (P < 0.05, ANOVA). dPGE caused a marked decrease in Kupffer cell and peritoneal macrophage TNF-alpha synthesis by 50 and 43%, respectively (P < 0.01, ANOVA). In conclusion, prostaglandins are critical for down regulating TNF-alpha production. Clinical use of cyclooxygenase inhibitors may result in adverse outcomes due to the excessive TNF-alpha production.

Burn injury and sepsis have been repeatedly demonstrated to impair the function of circulating (blood) neutrophils. As a result of the difficulty in harvesting and purifying neutrophils from the burn wound, there have been minimal investigations to date on the effect of burn injury and sepsis on the function of neutrophils which have reached the wound. We utilized a sponge matrix model in order to obtain neutrophils from burned and burned-infected rats. Despite having a higher concentration of neutrophils in the blood, both the burned and burned-infected rats were noted to have a decreased number of neutrophils infiltrating the sponge compared with the controls (1.91 +/- 0.30 x 10(6), 2.31 +/- 0.47 x 10(6), and 4.82 +/- 0.64 x 10(6) neutrophils per sponge, respectively). Blood neutrophils from both the burned and burned-infected rats had a greater chemiluminescence capacity than neutrophils from the control group (p < 0.0001). This enhanced capacity was not present with sponge neutrophils obtained from the burned-infected group. The diminished capacity may have been the result of a decreased concentration of prostaglandin E in the sponge fluid of the burned-infected rats compared with that of the burned or control rats (52 +/- 9, 135 +/- 15, and 114 +/- 13 pg/mL of sponge fluid, respectively).