[Diagnosis and treatment of blunt high-grade pancreatic trauma].

The clinical data of 20 patients with blunt high-grade pancreatic trauma who were admitted to the Department of Hepatobiliary and Pancreatic Surgery of Changhai Hospital Affiliated to Naval Military Medical University from December 2003 to February 2022 were retrospectively analyzed. There were 15 males and 5 females with a median age of 39 years (range: 14-54 years). The degree of pancreatic injury was graded according to the American Association for the Surgery of Trauma (AAST) scale, including 10 cases of grade Ⅲ (50%), 8 cases of grade Ⅳ (40%), and 2 cases of grade Ⅴ (10%). Then, the strategy of diagnosis and treatment for blunt high-grade pancreatic trauma was summarized. The diagnostic rate of CT was 78.9%. Finally, 17 cases (85%) were cured and 3 cases (15%) died. Among the 10 patients with grade Ⅲ pancreatic injury, 7 cases received distal pancreatectomy and splenectomy, 1 case received distal pancreatectomy with spleen preserved, 1 case received pancreatic duct stent placement under endoscopic retrograde cholangiopancreatography (ERCP) and percutaneous catheter drainage (PCD), and 1 case received only PCD. Among 8 cases of grade Ⅳ, 3 cases underwent Roux-en-Y pancreaticojejunostomy, 1 case received distal pancreatectomy and splenectomy, 1 case underwent distal pancreatectomy with spleen preserved, 2 cases received necrotic tissue removal+external drainage of pancreatic duct+abdominal drainage, and 1 case received exploratory laparotomy and gauze packing hemostasis. For 2 cases of grade Ⅴ, 1 underwent pylorus preserving pancreaticoduodenectomy, and the other case underwent pancreaticoduodenectomy combined with right hemicolectomy and splenectomy. Therefore, the treatment of blunt high-grade pancreatic trauma should follow the individualized treatment strategy, pay attention to the control of bleeding, extensive external drainage, appropriate debridement and resection and rational application of damage control surgery, select appropriate patients for conservative treatment, and ultimately benefit the patient.

[Diagnostic efficacy for predicting intraductal papillary mucinous neoplasms of the pancreas with high grade dysplasia or invasive carcinoma based on the surgery indications in different guidelines].

Objective: To evaluate the performance of the European Evidence-based Guidelines on Pancreatic Cystic Neoplasms (EEGPCN)(2018) and International Association of Pancreatology(IAP) Guideline(Version 2017) in predicting high grade dysplasia/invasive carcinoma-intraductal papillary mucinous neoplasm(HGD/INV-IPMN). Methods: A retrospective analysis of 363 patients,who underwent surgical resection in Changhai Hospital affiliated to Navy Medical University from January 2012 to December 2018 and were pathologically identified as (intraductal papillary mucinous neoplasm, IPMN),was performed. The patients,including 230 males and 133 females,aging (61.7±10.1) years(range:19 to 83 years). The proportion of HGD/INV-IPMN who met with the absolute indication(AI) of EEGPCN and high risk stigma(HRS) of IAP were compared. The binary Logistic regression analysis was used to find the independent risk factors of HGD/INV-IPMN.Eight combinations of risk factors derived from relative indication/worrisome feature or risk factors in this study,were made to evaluate the diagnostic efficacy. The area under curve(AUC) of receiver operating characteristics was used to evaluate the the cutoff value of risk factors(①CA19-9≥37 U/ml,②diameter of main pancreatic duct 5.0-9.9 mm,③enhancing mural nodule<5 mm,④(acute) pancreatiti,⑤acyst diameter ≥40 mm,⑤bcyst diameter ≥30 mm, ⑥thickened or enhancing cyst walls,⑦neutrophile granulocyte to lymphocyte ratio(NLR)≥2, ⑧cyst located in head, uncinate or neck,⑨carcinoembryonic antigen(CEA) ≥5 µg/L) number for predicting HGD/INV-IPMN.The accuracy,sensitivity,specificity,positive predictive value,negative predictive value,true positive,true negative,false positive,false negative,positive likelihood ratio,negative likelihood ratio,Youden index and F1 score were calculated. Results: Ninety-two patients(49.5%) of 186 ones who met AI and 85 patients(48.3%) of 176 ones who met HRS were respectively confirmed as HGD/INV-IPMN. In those patients who were not met AI,tumor location,thickened/enhancing cyst wall,CA19-9 elevated,NLR≥2 and CEA elevated were significantly (P<0.05) correlated with HGD/INV-IPMN. And tumor location(head/uncinate/neck vs. body/tail,OR=3.284,95%CI:1.268-8.503,P=0.014),thickened/enhancement cyst wall (with vs.without,OR=2.713,95%CI:1.177-6.252,P=0.019),CA19-9(≥37 U/L vs.<37 U/L, OR=5.086,95%CI:2.05-12.62,P<0.01) and NLR(≥2 vs.<2,OR=2.380,95%CI:1.043-5.434,P=0.039) were the independent risk factors of HGD/INV-IPMN. Patients with ≥4 risk factors of 9 in combination Ⅷ(①②③④⑤b⑥⑦⑧⑨) were diagnosed as HGD/INV-IPMN with the moderate accuracy(71.0%),moderate sensitivity (62.0%) and moderate specificity (73.0%). Patients with ≥4 risk factors of 9 in Combination Ⅶ(①②③④⑤a⑥⑦⑧⑨) were diagnosed as HGD/INV-IPMN with the highest specificity(83.0%) and patients with ≥3 risk factors of 8 in combination Ⅵ(①②③④⑤b⑥⑧⑨) were diagnosed as HGD/INV-IPMN with the highest sensitivity(74.0%). The AUC for diagnosis of HGD/INV-IPMN in combination Ⅵ,Ⅶ and Ⅷ were 0.72,0.75 and 0.75,respectively. Older patients and younger patients could respectively refer to combination Ⅶ and combination Ⅵ to improve the management of IPMN. Conclusions: Patients who meet AI of EEGPCN should undertake resection, otherwise the method we explored is recommended. The method of improvement for diagnosis of HGD/INV-IPMN is relatively applicable and efficient for decision-making of surgery, especially for younger patients with decreasing of missed diagnosis and elder patients with decreasing of misdiagnosis.

Impacts of COVID-19 on the electric vehicle industry: Evidence from China.

Electric vehicle development is critical to achieve the sustainable goals, while the hit of COVID-19 strikes the market and brings challenges to the whole industry. China, among one of the earliest regions affected by COVID-19 and takes a great part in the global electric vehicle market, is attracting growing attention on its post-pandemic trends in the electric vehicle industry. This paper provides a comprehensive analysis of COVID-19 impacts on China's electric vehicle industry from both the demand side and the supply side. Both challenges and opportunities for China's electric vehicle development are revealed with emerging trend analysis. It is found that the COVID-19 outbreak has reduced electric vehicle sales in the short-term, but may also stimulate future electric vehicle demand especially for large electric cars with better performance. Meanwhile, travel restrictions caused by COVID-19 have interrupted electric vehicle material supplies that relying on imports, accelerating domestic substitute exploitation and inventory improvement for critical parts. Additionally, massive lockdowns for controlling COVID-19 have disrupted productions and operations, which tends to expel small brands out of the competitive market, concentrating China's electric vehicle industry to the leading brands. Finally, the social distancing trend after pandemic is bringing challenges to traditional EV distribution channels with dealers, pushing automakers to develop innovative online selling channels. These impacts are likely to lead to a reformation of China's electric vehicle industry towards a more advanced and reliable future.

[Prognostic value of important driver gene mutations in patients with radical resection of pancreatic cancer].

Objective: To examine the prognostic value of four important driver gene mutations in patients with radical resection of pancreatic cancer. Methods: The clinical data and follow-up data of pancreatic cancer patients undergoing radical pancreatectomy and targeted sequencing from January 2016 to March 2018 at Department of Hepato-Biliary-Pancreatic Surgery, Changhai Hospital were retrospectively analyzed.There were 159 males and 88 females,aged of (60.8±8.7)years(range:33-83 years) and preoperative CA19-9 of (492.4±496.6)kU/L(range: 2-1 200 kU/L). One hundred and fifty nine cases of tumors were located in the head and 88 cases in the body and tail of the pancreas. After univariate analysis of clinical pathological factors (including gender, age, preoperative CA19-9, tumor location, tumor differentiation, pathological T and N stage, Micr. perineural invasion, Micr. lympho-vascular invasion, resection margin), the variable whose P<0.1 was included in COX regression model with four important driver gene mutations to find which mutation was related to prognosis independently. The number of gene mutations and KRAS subgroups were analyzed by Kaplan-Meier curve. Results: Among 247 patients,the number of KRAS,TP53, SMAD4 and CDKN2A mutations was 212 cases(85.8%), 160 cases(64.8%), 66 cases(26.7%) and 44 cases(17.8%),respectively.KRAS mutation was correlated with the tumor differentiation and pathological T stage (χ(2)=24.570/6.690, P=0.000/0.035), TP53 mutation was correlated with the tumor differentiation and the resected margin(χ(2)=5.500/4.620, P=0.019/0.032), and CDKN2A mutation was correlated with gender(χ(2)=16.574, P=0.000).COX regression model analysis showed that only KRAS mutation was an independent risk factor for disease free survival and overall survival(HR=1.776, 95%CI: 1.079-2.923, P=0.024; HR=1.923, 95%CI: 1.016-3.639, P=0.045); KRAS(G12D) mutation was associated with shorter OS(P=0.007). Conclusion: KRAS and its subgroup KRAS(G12D) mutation can be used as a prognostic index for patients with radical resection of pancreatic cancer.

The effect of dietary protein levels on the expression of genes coding for four selected protein translation initiation factors in muscle tissue of Wujin pig.

The objective of this study was to investigate the regulatory mechanism underlying the increased muscle protein accumulation in pigs while were fed a high protein diet. The eukaryotic initiation factors (eIFs) have been reported to involve in muscle protein synthesis. We investigated the mRNA and protein expression levels of eIF2B1, 4A1, 4B and 4E in Wujin pigs fed either a high protein (HP: 18%) or a low protein (LP: 14%) diet at 30, 60 or 100 kg body weight, based on real-time PCR and western blotting analyses. Our results indicated that the expression levels of eIF2B1 mRNA and protein were increased by HP diet at all body weight. The HP diet showed higher mRNA and protein levels of eIF4B gene at 60 and 100 kg. The protein expression of eIF4E phosphorylation was increased by HP diet only at 30 kg. These data suggested that the HP diet promoted porcine muscle protein accumulation mainly by up-regulating eIF2B1, 4B and 4E rather than 4A1 expression along the growth stages.

Expression of lipogenic genes during porcine intramuscular preadipocyte differentiation.

Intramuscular fat (IMF) content plays an important role in meat quality. Triglyceride (TG) metabolism in intramuscular adipocytes is strongly associated with the intramuscular fat deposition. To better understand the mechanisms leading to IMF deposition we compared the expression levels of genes related to preadipocyte differentiation and lipogenesis in the intramuscular preadipocytes isolated from the longissimus muscle of Wujin and Landrace pigs. The results showed that the intramuscular preadipocytes could differentiate into mature adipocytes in vitro. Triglyceride content in adipocytes isolated from Wujin pigs was higher than Landrace pigs during the middle and later phases of preadipocyte differentiation. The expression levels of genes related to preadipocyte differentiation such as PPARG and CEBPA showed differential expression between Wujin and Landrace porcine adipocytes during the early stage of differentiation. The expression levels of lipogenic genes such as FASN and SREBF1 were significantly higher in Wujin porcine intramuscular preadipocytes than in Landrace intramuscular preadipocytes at the middle and the later stages of differentiation. This suggests that preadipocyte differentiation and lipogenesis exhibited breed-related scheduling.

Molecular characterization and tissue expression profile of three novel ovine genes: ATP5O, NDUFA12 and UQCRH from muscle full-length cDNA library of black-boned sheep.

Three novel ovine genes were obtained from muscle full-length cDNA library of black-boned sheep. Sequence analysis revealed that nucleotide sequences of these genes were not homologous to any of the known sheep or goat genes, but these genes have high similarity to ATP synthase subunit O (ATP5O), NADH dehydrogenase (ubiquinone) 1 alpha subcomplex, 12 (NDUFA12) and ubiquinol-cytochrome c reductase hinge protein (UQCRH) genes of other mammal animals (accession number: FJ546085, FJ546078 and FJ546083). The alignment analysis showed that the ovine ATP5O, NDUFA12 and UQCRH genes and proteins have closer genetic relationships with the ATP5O, NDUFA12 and UQCRH genes and proteins from cattle. Conserved domain prediction showed that these three genes included OSCP, NDUFA12 superfamily and UCR-hinge superfamily domains respectively. The deduced sequence of ATP5O, NDUFA12 and UQCRH protein had 213, 145 and 91 amino acid residues, with a molecular weight of approximately 23419.66, 17089.50 and 10657.75 Da and a theoretical isoelectric point of 9.90, 9.68 and 4.45. The secondary structure prediction revealed that 60% helix structure in ATP5O, 60% coils in NDUFA12 and no strand in UQCRH. One potential signal peptide structure in ATP5O protein were found. NDUFA12 and UQCRH have the extremely low possibility of signal peptides. Meanwhile, RasMol was used for visualizing the PDB files generated by Swiss-Model in cartoon or three-dimensional format. ATP5O and UQCRH protein were modeled by Swiss-Model. Tissue expression profile indicated that the ovine ATP5O, NDUFA12 and UQCRH genes could be expressed in all detected tissues including muscles, heart, liver, spleen, lung, kidney and adipose tissues, but the expression abundance of these genes were various in the different tissues. Our experiment supplied the primary foundation for further researches on these three ovine genes.

Anti-adipocyte scFv-Fc antibody suppresses subcutaneous adipose tissue development and affects lipid metabolism in minipigs.

Anti-adipocyte monoclonal antibody has been shown to reduce body fat mass in animals. Here, we investigated the effects of an anti-adipocyte antibody (single-chain variable fragment and crystallizable fragment, scFv-Fc) on pig subcutaneous adipose tissue development and lipid metabolism. The scFv-Fc antibody did not alter feed intake or body weight of treated pigs. It suppressed subcutaneous adipose tissue development by reducing the percentage of larger adipocytes, which led to a reduction in body fat mass and subcutaneous adipose tissue thickness. Body fat mass was reduced by reducing triglyceride biosynthesis and promoting triglyceride lipolysis in adipose tissue. There was an increase in lipoprotein lipase mRNA expression in adipose tissue and activity in blood and an enhanced transportation of circulating high-density lipoprotein, low-density lipoprotein, and free fatty acids. Blood concentrations of triglyceride, total cholesterol, glucose, insulin, and adiponectin and mRNA expression of adiponectin in adipose tissue remained unaffected. These findings suggest that anti-adipocyte scFv-Fc antibody may have an application for reducing body fat mass in obese subjects.

Differential expression of lipid metabolism related genes in porcine muscle tissue leading to different intramuscular fat deposition.

Intramuscular fat (IMF) content affects meat quality and varies in different pig breeds. However, the underlying mechanisms of different IMF depositions in different genetic backgrounds of pigs have not been fully elucidated as yet. Lipid metabolism theoretically contributes to the variation of IMF content. The expression levels of genes and proteins as well as enzyme activities implicated in muscle lipid metabolism were investigated, which included lipogenetic genes (SREBP-1c and FAS), fatty acid transporting genes (H-FABP and A-FABP), fatty acid oxidative gene (CPT-1B) and lipolytic genes (ATGL and HSL) as well as the desaturated fatty acid gene (SCD). Longissimus muscle samples were collected from fatty Wujin pigs and lean Landrace pigs. Results showed that the average daily gain of Wujin pigs was lower than that of Landrace pigs. Wujin pigs had greater adipocyte diameter, IMF content and PUFA percentage than that of Landrace pigs. Compared with Landrace pigs, Wujin pigs exhibited higher expression levels, both in mRNA and protein, of FAS, SREBP-1c, SCD, A-FABP and H-FABP genes and lower expression levels of CPT-1B, HSL and ATGL genes. Overall, Wujin pigs possessed higher mRNA abundance, protein expression or enzyme activities of anabolism, fatty acid transportation and desaturation, and lower catabolism. Therefore, the mechanism of higher IMF content in fatty pigs may be due to the higher capacity of lipogenesis and fatty acid transportation and the lower capacity of lipolysis.

Effect of monoclonal antibody on expression of lipid metabolism related genes in porcine adipocytes.

In order to study the mechanism of monoclonal antibody (McAb) against a porcine 40-kDa adipocyte-specific plasma membrane protein in reducing fat deposition, porcine primary adipocytes were treated with the McAb during the process of adipocyte differentiation; its effect on expression of lipid metabolism related genes was investigated. Adipocytes were treated with 1-methyl-3-isobutylmethylxanthine (IDX) plus 10 microg/mL of the McAb or without McAb. The mRNA levels of adipocyte differentiation related genes (PPARgamma and C/EBPalpha), lipid metabolism related genes (FAS, HSL, CPT-1B, DGAT and A-FABP) and adiponectin gene (AdipoQ) were determined using real-time quantitative PCR. The results showed that the differentiated adipocyte number and triglyceride (TG) content in adipocytes treated with the McAb were lower than that in cells without McAb during the whole process of adipocyte differentiation. The McAb significantly reduced mRNA expression of PPARgamma, C/EBPalpha, FAS, DGAT, A-FABP and adiponectin genes, but increased mRNA expression of HSL and CPT-1B genes during the medium and latter stage of adipocyte differentiation. This suggested that the McAb decreased triglycerol accumulation in adipocyte by both inhibiting adipocyte differentiation and regulating lipid metabolism, especially at the medium and latter stage of porcine adipocyte differentiation.

Molecular cloning, sequence identification and tissue expression profile of three novel sheep (Ovis aries) genes - BCKDHA, NAGA and HEXA.

The complete coding sequences of three sheep genes- BCKDHA, NAGA and HEXA were amplified using the reverse transcriptase polymerase chain reaction (RT-PCR), based on the conserved sequence information of the mouse or other mammals. The nucleotide sequences of these three genes revealed that the sheep BCKDHA gene encodes a protein of 313 amino acids which has high homology with the BCKDHA gene that encodes a protein of 447 amino acids that has high homology with the Branched chain keto acid dehydrogenase El, alpha polypeptide (BCKDHA) of five species chimpanzee (93%), human (96%), crab-eating macaque (93%), bovine (98%) and mouse (91%). The sheep NAGA gene encodes a protein of 411 amino acids that has high homology with the alpha-N-acetylgalactosaminidase (NAGA) of five species human (85%), bovine (94%), mouse (91%), rat (83%) and chicken (74%). The sheep HEXA gene encodes a protein of 529 amino acids that has high homology with the hexosaminidase A(HEXA) of five species bovine (98%), human (84%), Bornean orangután (84%), rat (80%) and mouse (81%). Finally these three novel sheep genes were assigned to GenelDs: 100145857, 100145858 and 100145856. The phylogenetic tree analysis revealed that the sheep BCKDHA, NAGA, and HEXA all have closer genetic relationships to the BCKDHA, NAGA, and HEXA of bovine. Tissue expression profile analysis was also carried out and results revealed that sheep BCKDHA, NAGA and HEXA genes were differentially expressed in tissues including muscle, heart, liver, fat, kidney, lung, small and large intestine. Our experiment is the first to establish the primary foundation for further research on these three sheep genes.

Cloning and expression levels of genes relating to the ovulation rate of the Yunling black goat.

This study investigated the nucleotide sequences and tissue expression levels of genes relating to the ovulation rate in Yunling black goats, a famous Yunnan province, China, local breed with low fecundity. Five genes, FSHB, FSHR, BMP15, BMPR1B, and ESR2, were investigated; the complete cDNA sequences of these genes were 390, 2088, 1185, 1509, and 1585 bp, respectively, and compared with Boer goats (a more fecund breed), the sequence identities were 99%, 99%, 99%, 100%, and 99%, respectively. There were two base differences in FSHB and BMP15, four in FSHR, and three in ESR2. There were fewer follicles and oocytes in Yunling black goats than in Boer goats. Expression levels of FSHB, FSHR, and BMP15 genes in Yunling black goats were lower, and expression levels of BMPR1B and ESR2 genes were higher. Serum FSH content was lower in Yunling black goats, but serum estrogen content was higher. Protein expression levels of FSHR, BMP15, BMPR1B, and ESR2 in ovary and pituitary correlated positively with gene mRNA expression levels. In Yunling black goats, the mRNA expression levels of FSHB, FSHR, and BMP15 positively correlated with litter size, but those of BMPR1B and ESR2 correlated negatively. Together, base changes and variations in mRNA and protein expression levels of genes relating to the ovulation rate result in low fecundity in the Yunling black goat. Reduced BMP15 and FSHR levels may be related to the observed fewer oocytes and, consequently, fewer follicles.

Molecular cloning, sequence identification and tissue expression profile of three novel sheep (Ovis aries) genes - BCKDHA, NAGA and HEXA

The complete coding sequences of three sheep genes- BCKDHA, NAGA and HEXA were amplified using the reverse transcriptase polymerase chain reaction (RT-PCR), based on the conserved sequence information of the mouse or other mammals. The nucleotide sequences of these three genes revealed that the sheep BCKDHA gene encodes a protein of 313 amino acids which has high homology with the BCKDHA gene that encodes a protein of 447 amino acids that has high homology with the Branched chain keto acid dehydrogenase El, alpha polypeptide (BCKDHA) of five species chimpanzee (93 percent), human (96 percent), crab-eating macaque (93 percent), bovine (98 percent) and mouse (91 percent). The sheep NAGA gene encodes a protein of 411 amino acids that has high homology with the alpha-N-acetylgalactosaminidase (NAGA) of five species human (85 percent), bovine (94 percent), mouse (91 percent), rat (83 percent) and chicken (74 percent). The sheep HEXA gene encodes a protein of 529 amino acids that has high homology with the hexosaminidase A(HEXA) of five species bovine (98 percent), human (84 percent), Bornean orangután (84 percent), rat (80 percent) and mouse (81 percent). Finally these three novel sheep genes were assigned to GenelDs: 100145857, 100145858 and 100145856. The phylogenetic tree analysis revealed that the sheep BCKDHA, NAGA, and HEXA all have closer genetic relationships to the BCKDHA, NAGA, and HEXA of bovine. Tissue expression profile analysis was also carried out and results revealed that sheep BCKDHA, NAGA and HEXA genes were differentially expressed in tissues including muscle, heart, liver, fat, kidney, lung, small and large intestine. Our experiment is the first to establish the primary foundation for further research on these three sheep genes.

[Cloning, nucleotide sequence and tissue expression profile of three novel porcine genes--RHOB, RHOG and PRAF1].

The complete coding sequences of three porcine genes Rho-related GTP-binding proteins RHOB and RHOG and Prenylated Rab acceptor protein 1 (PRAF1) were amplified using the reverse transcriptase polymerase chain reaction based on the sequence information of the mouse or other mammals and referenced highly homologous pig ESTs. The nucleotide sequences of these three genes revealed that porcine RHOB gene encodes a protein of 196 amino acids that contains the conserved putative RhoA-like domain and has high homology with the RHOB precursor of human, rat and mouse (100%).The porcine RHOG gene encodes a protein of 191 amino acids that contains the conserved putative RhoG domain and has high homology with the RhoG precursor (RHOG) of human, mouse and Cricetus cricetus (98%). The porcine PRAF1 gene encodes a protein of 185 amino acids that contains the conserved putative PRA1 domain and has high homology with the PRAF1 of dog (97%), cattle (97%), human (96%), rat (95%) and mouse (95%). The tissue expression analysis indicated swine RHOB gene was moderately expressed in lung, weakly in fat, spleen, kidney, and almost not expressed in small intestine, large intestine, liver, muscle. The swine RHOG gene was over-expressed in small intestine, large intestine, liver, and muscle, moderately expressed in kidney, weakly in spleen, and almost not expressed in fat and lung. The swine PRAF1 gene was over-expressed in fat and spleen, moderately in lung and kidney, weakly in small intestine and large intestine, and almost not expressed in liver and muscle.

cDNA cloning and tissue expression analyses of the encoding regions for three novel porcine genes- MJD1, CDC42 and NECD.

The cDNAs for Machado-Joseph disease protein 1 homolog (MJD1), cell division control protein 42 homolog precursor(CDC42) and necdin (NECD) genes of pig were amplified using the reverse transcriptase polymerase chain reaction (RT-PCR) based on the conserved coding sequence information of the MJD1, CDC42, and NECD genes from mouse and other mammals and the referenced porcine EST sequence information. Tissue expression analysis showed the swine MJD1, CDC42, and NECD genes were obviously differentially expressed in different tissues including muscle, heart, liver, backfat, kidney, lung, small intestine, and large intestine. Our experiment established the primary foundation for further research on these three swine genes.

Molecular characterization of the encoding regions and tissue expression analyses for three novel porcine genes--HNRPA1, YIPF5 and UB2D2.

The complete encoding regions of three porcine genes--heterogeneous nuclear ribonucleoprotein A1 (HNRPA1), YIP1 family member 5 (YIPF5) and ubiquitin-conjugating enzyme E2 D2 (UB2D2) were amplified using the reverse transcriptase polymerase chain reaction (RT-PCR) based on the conserved encoding region information of the mouse or other mammals and the referenced highly homologous pig ESTs of these conserved encoding regions. These three novel porcine genes were assigned to GeneID: 768103, 768112, and 780418. The phylogenetic tree analysis revealed that the swine HNRPA1 has closer genetic relationships with the HNRPA1 of mouse and rhesus monkey, but the swine YIPF5 has a closer genetic relationship with the YIPF5 of cattle and the swine UB2D2 shows an evolutional model different with the UB2D2 of other five species. The tissue expression analysis indicated that the swine HNRPA1 gene was moderately expressed in fat, spleen and kidney, weakly expressed in muscle and lung, and hardly expressed in small intestine, large intestine and liver. The swine YIPF5 gene was moderately expressed in fat and spleen, and hardly expressed in small intestine, large intestine, liver, lung, muscle and kidney. The swine UB2D2 gene was weakly expressed in lung, and hardly expressed in small intestine, large intestine, liver, muscle, fat, spleen and kidney. Our experiment established the primary foundation for further research on these three swine genes.

Isolation, sequence identification and tissue expression distribution of three novel porcine genes--RAB14, S35A3 and ITM2A.

The complete coding sequences of three porcine genes--RAB14, S35A3 and ITM2A were amplified using the reverse transcriptase polymerase chain reaction (RT-PCR) based on the conserved sequence information of the mouse or other mammals. The nucleotide sequence analysis of these three genes revealed that porcine RAB14 gene encodes a protein of 215 amino acids that contains the conserved putative Ras-related protein Rab-14 domain and has high homology with the Ras-related protein Rab-14 (RAB14) of four species--human and mouse (99%) and rat (100%), dictyostelium discoideum (71%). The porcine S35A3 gene encodes a protein of 325 amino acids that contains the conserved putative nucleotide-sugar transporter domain and has high homology with the UDP-N-acetylglucosamine transporter (S35A3) of five species--cattle (98%), dog (97%), human (96%), mouse (95%) and rat (94%). The porcine ITM2A gene encodes a protein of 254 amino acids that contains the conserved putative BRICHOS domain and has high homology with the integral membrane protein 2A (ITM2A) of two species--human (89%), and mouse (88%). The tissue expression analysis indicated that the swine RAB14 gene was over-expressed in fat, lung, spleen, and kidney, moderately in large intestine, weakly in small intestine, and hardly expressed in muscle and liver. The swine S35A3 gene was moderately expressed in large intestine, fat, and spleen, weakly in liver and lung, and almost not expressed in muscle, small intestine, and liver. The swine ITM2A gene was over-expressed in fat and spleen, moderately in lung, weakly in muscle, and hardly expressed in liver, small intestine, large intestine, and kidney. Our experiment established the primary foundation for further research on these three swine genes.

Cytomegalovirus infection impairs the nitric oxide synthase pathway: role of asymmetric dimethylarginine in transplant arteriosclerosis.

BACKGROUND: We hypothesized that cytomegalovirus (CMV) may contribute to the vasculopathy observed in cardiac allograft recipients by impairing the endothelial nitric oxide synthase pathway. We focused on asymmetric dimethylarginine (ADMA, the endogenous inhibitor of nitric oxide synthase) as a potential mediator of the adverse vascular effect of CMV. METHODS AND RESULTS: Heart transplant recipients manifested elevated plasma ADMA levels compared with healthy control subjects. Transplant patients with CMV DNA-positive leukocytes had higher plasma ADMA concentrations and more extensive transplant arteriopathy (TA). Human microvascular endothelial cells infected with the CMV isolates elaborated more ADMA. The increase in ADMA was temporally associated with a reduction in the activity of dimethylarginine dimethylaminohydrolase (DDAH, the enzyme that metabolizes ADMA). Infected cultures showed high levels of oxidative stress with enhanced endothelial production of superoxide anion. CONCLUSIONS: CMV infection in human heart transplant recipients is associated with higher ADMA elevation and more severe TA. CMV infection in endothelial cells increases oxidative stress, impairs DDAH activity, and increases ADMA elaboration. CMV infection may contribute to endothelial dysfunction and TA by dysregulation of the endothelial nitric oxide synthase pathway.

Mild hyperhomocysteinemia is not associated with cardiac allograft coronary disease.

BACKGROUND: Hyperhomocysteinemia is an independent risk factor for coronary disease and elevated plasma homocysteine levels have been documented in heart transplant recipients. The aim of this study was to test the hypothesis that homocysteine levels are associated with presence or absence of transplant coronary artery disease. METHODS: Forty-three non-smoking adults were recruited, all of whom had received a heart transplant between 2 and 7 yr previously. All 43 had blood drawn for fasting homocysteine level on the day of presentation. All patients had undergone diagnostic coronary angiography within the past 6 months. RESULTS: For all patients, the average fasting plasma homocysteine level was 17.0+/-SD 6.6 micromol/L with a range from 6.0 to 36.9 micromol/L. Twenty-six patients (60%) had fasting plasma homocysteine levels above 15.0 micromol/L. On the basis of arteriography, patients were categorized as those with angiographically normal (n=22) or abnormal (n=21) coronary arteries. There was no difference in the mean plasma homocysteine level comparing patients with angiographically normal (17.2+/-SD 7.0 micromol/L) to those with abnormal (16.8+/-SD 6.2 micromol/L) coronary arteries. Plasma homocysteine levels increased with increasing plasma creatinine levels (r=0.63, p<0.0001) and with decreasing vitamin B6 levels (r=-0.56, p<0.0001). CONCLUSIONS: Mild hyperhomocysteinemia is a consistent finding among heart transplant recipients. This finding was not associated with transplant coronary artery disease in our patients. The combination of renal dysfunction and vitamin B6 deficiency may explain the unusual prevalence of hyperhomocysteinemia in heart transplant recipients.

Impact of prophylactic immediate posttransplant ganciclovir on development of transplant atherosclerosis: a post hoc analysis of a randomized, placebo-controlled study.

BACKGROUND: Coronary artery disease occurs in an accelerated fashion in the donor heart after heart transplantation (TxCAD), but the cause is poorly understood. The risk of developing TxCAD is increased by cytomegalovirus (CMV) infection and decreased by use of calcium blockers. Our group observed that prophylactic administration of ganciclovir early after heart transplantation inhibited CMV illness, and we now propose to determine whether this therapy also prevents TxCAD. METHODS AND RESULTS: One hundred forty-nine consecutive patients (131 men and 18 women aged 48+/-13 years) were randomized to receive either ganciclovir or placebo during the initial 28 days after heart transplantation. Immunosuppression consisted of muromonab-CD3 (OKT-3) prophylaxis and maintenance with cyclosporine, prednisone, and azathioprine. Mean follow-up time was 4.7+/-1.3 years. In a post hoc analysis of this trial designed to assess efficacy of ganciclovir for prevention of CMV disease, we compared the actuarial incidence of TxCAD, defined by annual angiography as the presence of any stenosis. Because calcium blockers have been shown to prevent TxCAD, we analyzed the results by stratifying patients according to use of calcium blockers. TxCAD could not be evaluated in 28 patients because of early death or limited follow-up. Among the evaluable patients, actuarial incidence of TxCAD at follow-up (mean, 4.7 years) in ganciclovir-treated patients (n=62) compared with placebo (n=59) was 43+/-8% versus 60+/-10% (P<0.1). By Cox multivariate analysis, independent predictors of TxCAD were donor age >40 years (relative risk, 2.7; CI, 1.3 to 5.5; P<0.01) and no ganciclovir (relative risk, 2.1; CI, 1.1 to 5.3; P=0.04). Stratification on the basis of calcium blocker use revealed differences in TxCAD incidence when ganciclovir and placebo were compared: no calcium blockers (n=53), 32+/-11% (n=28) for ganciclovir versus 62+/-16% (n=25) for placebo (P<0.03); calcium blockers (n=68), 50+/-14% (n=33) for ganciclovir versus 45+/-12% (n=35) for placebo (P=NS). CONCLUSIONS: TxCAD incidence appears to be lower in patients treated with ganciclovir who are not treated with calcium blockers. Given the limitations imposed by post hoc analysis, a randomized clinical trial is required to address this issue.