[Risk factors for delayed gastric emptying after laparoscopic pancreaticoduodenectomy: a single-center experience of 1 000 cases].

Objective: To explore the causes and summarize the treatment experience for clinically relevant delayed gastric emptying(DGE) after laparoscopic pancreaticoduodenectomy(LPD). Methods: The clinical data of 1 000 patients who underwent LPD in the Department of Liver Transplantation and Hepatobiliary Surgery,Shandong Provincial Hospital Affiliated to Shandong First Medical University between March 2017 and September 2022 was retrospectively collected. There were 640 males and 360 females,with an age of (60.1±11.4)years(range: 13 to 93 years),and 590 patients were older than 60 years. Depending on the severity of DGE,patients were divided into a clinically relevant DGE group and a 0/A grade DGE group. The comparison between the two groups was performed by the χ2 test,Fisher's exact probability method,t test or the rank sum test,and the effects of various treatment strategies for clinically relevant DGE were evaluated. Results: LPD was conducted successfully in all 1 000 patients,with a surgical time of (344.8±103.6)minutes(range:160 to 450 minutes) and intraoperative blood loss (M(IQR)) of 100 (150) ml(range:50 to 1 000 ml). A total of 74 patients(7.4%) developed clinically relevant DGE. Compared to those in the 0/A grade DGE group,patients in the clinically relevant DGE group had a higher preoperative body mass index of ((24.9±3.5)kg/m2 vs. (23.9±3.3)kg/m2,t=-2.419,P=0.016),more postoperative bile leakage(51.4%(38/74) vs. 10.8%(100/926)),pancreatic fistula(59.5%(44/74) vs. 22.9%(212/926)),abdominal infection(74.3%(55/74) vs.14.6%(135/926)),and abdominal bleeding(43.2%(32/74) vs. 11.3%(105/926))(all P<0.05). Among these patients,10 cases(13.5%) received enteral nutrition treatment,22 cases(29.7%) received parenteral nutrition treatment,and 42 cases(56.8%) received a combination of enteral and parenteral nutrition treatment. The time for patients to return to a normal diet was 21(14)days (range: 8 to 85 days). Compared to those who received only enteral(23.5(27.0)days) or parenteral nutrition treatment(15.5(11.0)days),patients who received a combination of enteral and parenteral nutrition treatment(25.5(31.0)days) had a longer time to return to a normal diet (Z=20.019,P<0.01). Among the 60 patients who developed secondary DGE,48 cases(80.0%) received ultrasound-guided puncture and drainage treatment,while 12 cases(20.0%) only received anti-infection treatment. The patients in the non-puncture drainage group had a longer time to return to a normal diet than those in the puncture drainage group (26.5(12.5)days vs. 20.0(11.0)days, Z=-2.369,P=0.018). Conclusions: Patients with clinically relevant DGE after LPD had a higher proportion of postoperative complications such as pancreatic fistula,biliary fistula and abdominal infection. A combination of enteral and parenteral nutrition treatment is needed for patients with a long-term course of DGE."Smooth" drainage and ani-infectious therapy could contribute to the recovery of DGE.

[Advances in internal fixation for the treatment of extra-articular distal tibial fracture].

Extra-articular distal tibial fractures as a result of high-energy damage are often comminuted or displaced, frequently accompanied by severe soft tissue injuries.Poor blood supply and various complications make the treatment more difficult,affecting life quality of the patients.The main goals of the treatment are to abtain a healed,well-aligned fracture,functional range of motion of the ankle joint and minimizing complications.It is generally recommended that surgical treatment be performed in the proper context of local conditions to facilitate early functional exercise.Plate fixation and intramedullary nail fixation are the common options for closed fractures.This article focuses on the two treatment methods and some important auxiliary technologies in both domestic and foreign, hoping to provide some references for clinical treatment.

Immunogenicity of the envelope GP3 protein of porcine reproductive and respiratory syndrome virus displayed on baculovirus.

Porcine reproductive and respiratory syndrome virus (PRRSV) has been recognized as one of the most important pathogens of pigs throughout the world. The minor envelope protein GP3 of PRRSV plays an important role in clearing of the virus infection and protecting the animals. In this study, a recombinant baculovirus (BacSC-GP3) expressing His6-tagged GP3 with the transmembrane (TM) and cytoplasmic (CT) domains of envelope protein gp64 was constructed and its immunogenicity was evaluated in mouse and piglet models. The His6-tagged GP3 was successfully displayed on the surface of virions as well as virus-infected Sf-9 cells. The animals immunized with BacSC-GP3 gave a slightly higher (piglets) up to a markedly higher (mice) humoral and lymphocyte proliferation responses than those that received a commercial killed vaccine. This is the first study on the immunogenicity of recombinant GP3-baculovirus, which indicates that the latter can represent an alternative strategy for developing a more effective PRRSV vaccine.

[Expression of a snake venom plasminogen activator TSV-PA in insect cells].

A snake venom gene TSV-PA was inserted into the donor plasmids pFastBacHTa and expressed in Tn-5B1-4 cells. SDS-PAGE analysis revealed that the molecular weight of expressed product of TSV-PA were 33 kD. It was also proved by Western blot. The result of enzyme activity showed that TSV-PA protein expressed in insect cells had a higher activity.

Co-regulation of tissue-specific alternative human carnitine palmitoyltransferase Ibeta gene promoters by fatty acid enzyme substrate.

Carnitine palmitoyltransferase I (CPT-I) catalyzes the rate-determining step in mitochondrial fatty acid beta-oxidation. CPT-I has two structural genes (alpha and beta) that are differentially expressed among tissues. Our CPT-Ibeta isolates from a human cardiac cDNA library contained two different extreme 5'-sequences derived from short alternative first untranslated exons that utilize a common splice acceptor site in exon 2. Primer extension identified single dominant start sites for each transcript, and ribonuclease protection assays showed the presence of one 5'-exon in liver, muscle, and heart mRNAs, indicating that the cognate promoter U (upstream/ubiquitous) is active in each of these tissues. By contrast, mRNAs containing the alternative 5'-exon were present only in muscle and heart, indicating a muscle-specific promoter M (muscle). CPT-Ibeta mRNA levels increased markedly in tissues of fasted rats, when circulating free fatty acid concentrations are elevated. Using CPT-Ibeta promoter/reporter transient transfection of murine C2C12 myotubes and HepG2 hepatocytes, fatty acids were found to increase promoter activity in a peroxisome proliferator-activated receptor alpha (PPARalpha)-dependent fashion. A promoter fatty acid response element (FARE) was mapped, mutation of which ablated fatty acid-mediated production of both transcripts. PPARalpha/retinoid X receptor alpha formed specific complexes with oligonucleotides containing the FARE, and anti-PPARalpha antibody shifted nuclear protein-DNA complexes, confirming the role of this factor in regulating the expression of this critical metabolic enzyme gene. The constitutive repressor chicken ovalbumin upstream promoter transcription factor competitively binds at the FARE and modulates fatty acid induction of the promoters.

Expression of novel isoforms of carnitine palmitoyltransferase I (CPT-1) generated by alternative splicing of the CPT-ibeta gene.

Carnitine palmitoyltransferase I (CPT-I) catalyses the rate-determining step in mitochondrial fatty acid beta-oxidation. The enzyme has two cognate structural genes that are preferentially expressed in liver (alpha) or fat and muscle (beta). We hypothesized the existence of additional isoforms in heart to account for unique kinetic characteristics of enzyme activity in this tissue. Hybridization and PCR screening of a human cardiac cDNA library revealed the expression of two novel CPT-I isoforms generated by alternative splicing of the CPT-Ibeta transcript, in addition to the beta and alpha cDNA species previously described. Ribonuclease protection and reverse transcriptase-mediated PCR assays confirmed the presence of mRNA species of each splicing variant in heart, skeletal muscle and liver, with differing relative concentrations in the tissues. The novel splicing variants omit exons or utilize a cryptic splice donor site within an exon. Deduced polypeptide sequences of the novel enzymes include omissions in the region of putative membrane-spanning and malonyl-CoA regulatory domains compared with the previously described CPT-Is, implying that the encoded enzymes will exhibit unique features with respect to outer mitochondrial membrane topology and response to physiological and pharmacological inhibitors.

Rat carnitine palmitoyltransferase Ibeta mRNA splicing isoforms.

Carnitine palmitoyltransferase I (CPT-I) catalyzes the rate-determining step in mitochondrial fatty acid beta-oxidation. The enzyme has two cognate structural genes (alpha and beta) that are differentially expressed in tissues. We show multiple mature mRNAs in rat heart derived from alternative splicing of CPT-Ibeta transcripts. Two novel messages are deleted for regions of the previously described mRNA that encode membrane-spanning and regulatory domains, suggesting that the cognate isozymes will exhibit unique kinetic characteristics.

Inhibition of beta-amyloid cytotoxicity by midkine.

Midkine (MK) is a neurotrophic and angiogenic growth factor whose expression occurs mainly in fetus. It was reported that MK was present in senile plaques of Alzheimer's disease (AD). To investigate the role of MK during amyloid plaques formation in AD, we examined the in vitro effect of MK on Abeta aggregation and Abeta-induced cytotoxicity. We found that incubation of MK with Abeta resulted in the formation of MK/Abeta complexes. The C-terminus of MK (60-121) played a similar role as the full length MK in complex formation. This interaction of MK and Abeta demonstrated significant inhibition on Abeta self-aggregation. MK also inhibited the cytotoxicity of Abeta on PC12h cells. These findings suggest that MK protects the cells from Abeta-induced cytotoxicity through its complex formation with Abeta. MK is probably expressed to prevent cell death in AD.

Synchronous primary neuroendocrine carcinoma and adenocarcinoma in Barrett's esophagus.

We report a case of a synchronous primary small-cell carcinoma and adenocarcinoma of the esophagus with ectopic gastrin and calcitonin production. The double primary carcinomas appeared to arise from dysplastic Barrett's mucosa in a 43-year-old woman with a long history of reflux esophagitis. She is one of the few reported long-term survivors treated by surgical resection and combination chemotherapy. The histopathology and treatment of this rare tumor are briefly discussed.

Desmoid tumor of the sternum presenting as an anterior mediastinal mass.

A 20-year-old man with a large, asymptomatic mediastinal mass was found to have desmoid-type fibromatosis (DF) by needle biopsy. The tumor arose from the internal periosteum of the sternum and mimicked an anterior mediastinal mass. A wide resection of the sternum, including portions of the clavicles and costal cartilages, and reconstruction with a Gore-Tex soft tissue patch were performed. Although extremely rare, desmoid tumor of the sternum should be considered in the differential diagnosis of anterior mediastinal tumors.

[A sensitive fluorescent assay for the detection and quantification of mitochondrial 3243 mutation].

A mutation at nucleotide 3243 in the mitochondrial DNA (mtDNA) specifying tRNALeu(UUR) has been shown to be related with diabetes mellitus. Since mtDNA shows heteroplasmy and its composition would change among various tissues, it is important to quantitate the percentage of mutant mtDNA precisely. Here we report a rapid and sensitive method to determine the mutant mtDNA. A fragment of mtDNA containing nucleotide 3243 was amplified by PCR using a rhodamine-labeled primer and the product was digested with ApaI. The PCR product from normal gene was resistant to ApaI digestion, while a mutation from A to G generated an ApaI site in the mutant mtDNA and its PCR product was cleaved into two fragments. The digested product was separated by acrylamide gel electrophoresis and each product was quantitated by a fluorescence analyzer. PCR analysis using standard mixtures of normal and mutant mtDNAs indicated that this method can be used to detect as little as 1% mutant mtDNA.

Analysis of the thromboxane/prostacyclin balance in human urine by gas chromatography/selected ion monitoring: abnormalities in diabetics.

We microanalyzed 2,3-dinor-6-keto-prostaglandin F1 alpha (2,3-dinor-6-keto-PGF1 alpha 1) and 11-dehydrothromboxane B2 (11-dehydro-TXB2, 2) in human urine. Samples containing a [2H4]-analogue as an internal standard were extracted by chromatography using Sep Pak tC18 and silica gel. The compounds were then analysed by means of the lactone ring opening reaction and dimethylisopropylsilylation. The conversion of 1 to 1-methyl ester (ME)-propylamide (PA)-9, 12, 15-dimethylisopropylsilyl (DMIPS) ether derivative and of 2 to 1-ME-6-methoxime (MO)-9, 12, 15-tris-DMIPS ether derivative was followed by gas chromatography/selected ion monitoring (GC/SIM). Interfering substances from the urine matrix were eliminated during GC/SIM analysis using a DB-5 column. We were able to detect 1 (222-1031 pg/mg creatinine) and 2 (18-155 pg/mg creatinine) in human urine. Furthermore, the thromboxane/prostacyclin (IX/PGI) ratio in the urine of diabetics was higher than that of healthy volunteers. This method can be used to determine the TX/PGI balance in human urine.

Modulation of myocardial alpha 1- but not beta-adrenoceptors after 90-day tail-suspension.

We have previously demonstrated that prolonged simulated microgravity (tail-suspension) leads to cardiac alterations with increased resting heart rate, myocardial degradation changes and attenuated myocardial contractility. The present study investigated the potential role of adrenoceptor mechanisms underlying them. Changes of myocardial alpha 1-adrenoceptor (alpha 1-AR) and beta 1-adrenoceptor (beta-AR) in 90-day tail-suspended rats was investigated by the method of radioligand binding assay and application of Scatchard's method. The results showed significantly decreased quantity of specific binding of 125I-BE[2-beta-(4-hydroxy-3-[125I]indophenyl)-ethylaminomethyltetralone] to alpha 1-AR present in membrane derived from ventricular myocardium of the suspended animals, despite the affinity of the alpha 1-AR to 125I-Be was unchanged. But neither the quantity nor the affinity of beta-AR binding to 125I-Pindolol was significantly altered. In addition, the spontaneously beating rate of isolated right atria from tail-suspended animals showed little change in sensitivity and reactivity to the stimulations of graded phenylephrine (alpha-agonist, measured in the presence of beta-antagonist propranolol) and isoproterenol (beta-agonist), compared with the control rats. There were also no obvious differences of the effects of the isoproterenol on the contractility of isolated left ventricular papillary muscles between the two groups. Since myocardial alpha 1-AR mediated-effects include production of cardiac hypertrophy and enhancement of myocardial glucose uptake and glycolysis, the down-regulation of the alpha 1-AR may be a contributor to the cardiac cellular accumulation and the myocardial degradation changes as found in our tail-suspended rats. The data from this study also suggest that the myocardial beta-adrenoceptors are not affected by the prolonged tail-suspension.

Alterations of subtypes of cardiac adrenoceptors in old rat.

AIM: To determine alterations of subtypes of myocardial adrenoceptors in senescence. METHODS: Heart membrane preparations were made from 3- and 25-month old Wistar rats. Alpha 1- and beta-adrenoceptors were measured by radioligand, 125I-BE2254 and 125I-pindolol, binding assays, respectively. RESULTS: In the old rat heart, alpha 1- and beta-adrenoceptor densities were declined from the young rats of 119 +/- 4 and 45.9 +/- 1.9 pmol L-1 to 70 +/- 6 and 36.4 +/- 1.6 pmol L-1 (P < 0.01), with a greater change in alpha 1-AR and in beta-AR. The ratio of alpha 1A/alpha 1B subtypes was decreased from the young rats of 39/61 to the old rats of 26/74 (P < 0.05). CONCLUSION: The cardiac adrenoceptors are decreased with different extents in the different subtypes in old rats.

[Effects of long-term atenolol treatment on beta-adrenoceptor subtypes in rat heart].

The effects of long-term beta 1-AR selective antagonist atenolol treatment on beta-adrenoceptor subtypes were studied by radioligand binding assay, function determination and cAMP accumulation measerment in rat heart. The reasults showed that during long-term administration of atenolol: (1) The density of total beta-AR was increased by approximately 57%; the positive inotropic response and cAMP formation induced by activation of beta-AR were also enhanced. (2) The 125I-pindol competitive inhibition curve for CGP20712A showed that there were no significant difference in the percentage of beta 1- and beta 2-AR sites between the atenolol treated rats and the control rats; pA2 values for selective beta 1-AR antagonist CGP20712A and pKB values for selective beta 1-AR antagonist ICI 118, 551 were not significantly different in the two groups. The results suggested that beta 1- and beta 2-adrenoceptors were upregulated not only in density but also in positive inotropic effect to the same extent.

Role of alpha 1A- and alpha 1B-adrenoceptors in phenylephrine-induced positive inotropic response in isolated rat left atrium.

We determined the distribution of alpha 1A- and alpha 1B-adrenoceptor subtypes and their functional roles in phenylephrine (PE)-induced positive inotropic responses in rat atrium. Radioligand binding assays in membrane preparations of rat atria showed that 62% of [125I]BE 2254 binding sites were irreversibly inactivated by pretreatment with 20 microM chloroethylclonidine (CEC). Inhibition curves for WB 4101 and 5-methyl-urapidil (5-MU) were better fit by a two-site model, comprising 29-35% high-affinity sites and 65-71% low-affinity sites, suggesting that rat atria contains both alpha 1A and alpha 1B subtypes in a ratio of approximately 1:2. In isolated perfused atria, pretreatment with CEC inhibited the maximum PE-induced positive inotropic response by 55%, and pA2 values for WB 4101 and 5-MU in inhibiting this response were 8.26 +/- 0.4 and 7.85 +/- 0.07, respectively, between the KD values for alpha 1A and alpha 1B subtypes. PE shifted the concentration-contractile response curve for Ca2+ to the left and upward. Pretreatment with CEC completely abolished whereas 1 nM WB 4101 did not alter, the effect of PE on Ca2+ sensitivity. These results demonstrate that both alpha 1A- and alpha 1B- adrenoceptor subtypes are involved in the PE-induced positive inotropic response. Only activation of the alpha 1B subtype potentiates the positive inotropic effect induced by increasing extracellular Ca2+, however, which suggests that the mechanisms involved in the action of the two subtypes may differ at least in part.

[Effect of alpha 1-adrenoceptor subtypes on beta-adrenoceptor mediated positive inotropic response in rat left atria].

The distribution of the alpha 1-adrenoceptor (alpha 1-AR) subtypes and the effects of activation of alpha 1-AR subtypes on the beta-adrenoceptor (beta-AR) mediated positive inotropic response were investigated. The radioligand binding assays indicated that the Bmax and Kd values were 11.7 +/- 18 fmol/mg.protein and 86.0 +/- 9.6 pmol/L respectively. Pretreatment of the preparations with 20 mumol/L chloroethylclonidine (CEC) which inactivated alpha 1B subtype, decreased the Bmax to 45.7 +/- 5.2 fmol/mg.protein (P < 0.01). The inhibition curves of 5-methyl-urapidil were best fitted to two site model and indicated that alpha 1A subtype took 28.5% of total 125IBE specific binding sites. In the functional experiments, norepinephrine (NE) induced a positive inotropic response in a concentration dependent manner by activation of both beta- and alpha 1-AR. The concentration-response curves (CRC) for NE were shifted rightward after the pretreatment of the preparations with 20 mumol/L CEC, but leftward in the presence of 1 nmol/L WB4101. In the presence of 10 mumol/L phentolamine which inactivated both alpha 1-AR subtypes, the CRC for NE were shifted leftward. When alpha 1-AR was activated by phenylephrine the CRC for isoproterenol (selective beta-AR agonist) were shifted rightward. The results suggested that the alpha 1B subtype enhanced while the alpha 1A subtype inhibited the beta-AR mediated positive inotropic response. When both alpha 1A and alpha 1B subtypes were activated simultaneously the alpha 1A subtype showed a dominate role.

Microdetermination of 2,3-dinor-6-ketoprostaglandin F1 alpha in human urine using gas chromatography-high-resolution selected-ion monitoring.

The microdetermination of 2,3-dinor-6-ketoprostaglandin F1 alpha (I) in human urine is described. Samples to which the [2H4]-analogue was added as an internal standard were extracted by chromatographic sample preparation using a Bond Elut C18 cartridge and a silica gel column. Conversion of the extracted I into the 1-methyl ester-6-methoxime-9,11,15-trisdimethylisopropylsilyl ether derivative was followed by gas chromatography-high-resolution selected-ion monitoring (GC-HR-SIM). Interfering substances from the urine matrix were eliminated during GC-HR-SIM analysis using a DB-1 column. A good linear response over the range 10 pg-100 ng per tube was demonstrated. Compound I could be detected in the range 26-375 pg/ml of human urine. The proposed method can be applied to the determination of I in human urine.