Dietary soluble non-starch polysaccharide level and composition influences grower and finisher phase performance, excreta moisture content and total tract nutrient digestibility in broilers.

1. The aim of this study was to examine the impact of dietary soluble non-starch polysaccharide (sNSP) level and composition on grower and finisher phase performance, total tract nutrient digestibility and excreta moisture content in broiler chickens.2. Cobb 500 broilers (n = 1080) were fed 12 dietary treatments; four diets with differing primary grain sources (barley, corn, sorghum and wheat) and three different sNSP levels (low, medium and high). Diets were formulated to have similar protein and energy levels but differing sNSP levels, induced by manipulating the quantity of the ingredients in the diet. The diets were fed in three phases, starter (d 0-12), grower (d 12-23) and finisher (d 23-31).3. For birds aged d 23 and 31, total pen body weight and feed intake were determined, and fresh excreta and litter samples were collected per pen. Dry matter (DM) content was measured in the excreta and litter samples. Total tract DM digestibility, apparent metabolisable energy corrected to nitrogen (AMEn), and soluble and insoluble NSP and free oligosaccharide degradability were evaluated.4. In birds fed the sorghum- and corn-based diets, feeding high sNSP resulted in a lower cFCR at d 0-23 compared to low sNSP (P = 0.004 and P = 0.044, respectively). In birds fed the corn-based diet, feeding low sNSP resulted in the lowest litter DM but highest DM digestibility at d 23 (P = 0.045 and P < 0.001) and d 31 (P = 0.022 and P = 0.008). For all diets, degradability of sNSP was higher and insoluble NSP was lower when feeding low compared to high sNSP (P < 0.001). In birds fed the barley- and sorghum-based diets, AMEn was lower when feeding the low compared to high sNSP level (P < 0.001 and P = 0.016, respectively).6. Results from this study showed that level of dietary sNSP impacts broiler productive performance and nutrient utilisation.

Physico-chemical properties of late-incubation egg amniotic fluid and a potential in ovo feed supplement.

OBJECTIVE: This study explored the physico-chemical properties of late-incubation egg amniotic fluid and a potential in ovo feed (IOF) supplement. METHODS: Amniotic fluid was collected from broiler breeders (Ross 308, 51 weeks and Cobb 500, 35 weeks) on day 17 after incubation. A mixture of high-quality soy protein supplement - Hamlet Protein AviStart (HPA) was serially diluted in MilliQ water to obtain solutions ranging from 150 to 9.375 mg/mL. The mixtures were heat-treated (0, 30, 60 minutes) in a waterbath (80°C) and then centrifuged to obtain supernatants. The amniotic fluid and HPA supernatants were analysed for their physico-chemical properties. RESULTS: Only viscosity and K+ were significantly (p<0.05) different in both strains. Of all essential amino acids, leucine and lysine were in the highest concentration in both strains. The osmolality, viscosity and pCO2 of the supernatants decreased (p<0.05) with decreasing HPA concentration. Heat treatment significantly (p<0.05) affected osmolality, pH, and pCO2, of the supernatants. The interactions between HPA concentration and heat treatment were significant with regards to osmolality (p<0.01), pH (p<0.01), pCO2 (p<0.05), glucose (p<0.05), lactate (p<0.01) and acid-base status (p<0.01) of HPA solutions. The Ca2+, K+, glucose, and lactate increased with increasing concentration of HPA solution. The protein content of HPA solutions decreased (p<0.05) with reduced HPA solution concentrations. The supernatant from 150 mg/mL HPA solution was richest in glutamic acid, aspartic acid, arginine and lysine. Amino acids concentrations were reduced (p<0.05) with each serial dilution but increased with longer heating. CONCLUSION: The values obtained in the primary solution (highest concentration) are close to the profiles of high-protein ingredients. This supplement, as a solution, hence, may be suitable for use as an IOF supplement and should be tested for this potential.

Incidence of Hyperglycemia during Induction of Remission Phase of Pediatric Acute Lymphoblastic Leukemia.

Hyperglycemia is frequently associated with acute lymphoblastic leukemia (ALL) therapy. It has long been recognized as a consequence of corticosteroids and asparaginase. These medications are usually administrated concurrently in high doses during the initial induction phase of chemotherapy. As a result, hyperglycemia frequently develops during this phase, with resolution after the steroids and asparaginase have been discontinued or reduced in dose. Various recent studies have evaluated the incidence of hyperglycemia in this particular group of patients and its relationship with chemotherapy. However the incidence of hyperglycemia during induction of remission therapy in children with ALL is not known, and is the subject of this study. To determine the incidence as well as identification of day of onset and duration of hyperglycemia and other complications during induction of remission therapy. This prospective cross-sectional study included 50 newly diagnosed patients aged 1-15 years and was done from December 2010 to May 2011 in the department of Pediatric Haemato Oncology, Bangabandhu Sheikh Mujib Medical University, Bangladesh. Hyperglycemia was defined as ≥2 random glucose determinations of ≥200mg/dl during the first 28 days of induction chemotherapy. Out of 50 patients, 4(8%) patients developed hyperglycemia during remission induction therapy. No significant difference was found between two groups regarding age distribution (p>0.05) and body weight (p>0.05). Among patients who developed hyperglycemia 3 patients (75%) and 1 patient (25%) experienced hyperglycemia during second week and third week of induction therapy respectively. Hyperglycemia persists <7 days in case of 3(75%) patients and >7 days in case of 1(25%) patients. The incidence of chemotherapy induced hyperglycemia in this present study is 8%. Most patients who develop therapy induced hyperglycemia recover when L-asparaginase and corticosteroids are discontinued and they suffer no long term adverse effects.

Energy Value of Cassava Products in Broiler Chicken Diets with or without Enzyme Supplementation.

This study investigated the metabolizable energy (ME) intake, net energy of production (NEp), heat production (HP), efficiencies of ME use for energy, lipid and protein retention as well as the performance of broiler chickens fed diets based on cassava chips or pellets with or without supplementation with an enzyme product containing xylanase, amylase, protease and phytase. The two products, cassava chips and pellets, were analysed for nutrient composition prior to feed formulation. The cassava chips and pellets contained 2.2% and 2.1% crude protein; 1.2% and 1.5% crude fat; and 75.1% and 67.8% starch, respectively. Lysine and methionine were 0.077%, 0.075%, and 0.017%, 0.020% protein material, respectively, while calculated ME was 12.6 and 11.7 MJ/kg, respectively. Feed intake to day 21 was lower (p<0.01) on the diet containing cassava chips compared to diets with cassava pellets. Enzyme supplementation increased (p<0.01) feed intake on all diets. Live weight at day 21 was significantly (p<0.01) reduced on the diet based on cassava chips compared to pellets, but an improvement (p<0.01) was noticed with the enzyme supplementation. Metabolizable energy intake was reduced (p<0.01) by both cassava chips and pellets, but was increased (p<0.01) on all diets by enzyme supplementation. The NEp was higher (p<0.01) in the maize-based diets than the diets containing cassava. Enzyme supplementation improved (p<0.01) NEp in all the diets. Heat production was highest (p<0.01) on diets containing cassava pellets than on cassava chips. It is possible to use cassava pellets in diets for broiler chickens at a level close to 50% of the diet to reduce cost of production, and the nutritive value of such diets can be improved through supplementation of enzyme products containing carbohydrases, protease, and phytase.

Evaluation of extra pulmonary tuberculosis in Bangladeshi patients.

This is a retrospective study of 200 cases of Extra Pulmonary Tuberculosis (EPTB) and this study was conducted from January 2010 to July 2011 at Sheheed Ziaur Rahman Medical College Hospital and TB clinic, Bogra, Bangladesh. Data were collected from the Hospital records on demographics, clinical, laboratory and treatment out come status. The incidence of EPTB was high in 16-45 years age group (55%), mean age was 35.67±14.6 years and predominated in Female patients (60%). Extra pulmonary Tuberculosis (EPTB) still constitutes an important clinical problem in Bangladesh. The objective of this study was to evaluate the demography and features of patients with EPTB in our high burden tuberculosis country. Lymph nodes are the most common site of involvement (50%) followed by tubercular pleural effusion (15%) and virtually every site of the body can be affected by tuberculosis. Since the clinical presentation of EPTB is atypical, tissue samples for the confirmation of diagnosis can sometimes be difficult procedure and conventional diagnostic method have a poor yield, so the diagnosis is often delayed. EPTB constitutes about 15-20% of all cases of TB patients and it is more common in low socio-economic group (60%) Biopsy and/ or surgery, FNAC is required to procure tissue samples and pus and/or aspirated fluids are required for diagnosis and for managing complications. The EPTB usually responds to standard anti tubercular drug regimen.

Urinary microalbumin as a risk factor for ischaemic stroke.

Microalbuminuria (MA), a microvascular irregularity of the blood - urine interface within kidney glomeruli, may reflect the renal sign of global endothelial dysfunction, and has been associated with an elevated risk of serious cardiovascular events including stroke. This case-control study was carried out in the Department of Neurology, Bangabandhu Sheikh Mujib Medical University (BSMMU), Dhaka, Bangladesh, from January 2008 to December 2009. This relationship appears graded with higher levels of microalbuminuria being independently associated with a greater magnitude of vascular risk over time. We studied 60 patients of ischemic stroke with in 4 weeks of onset of symptoms and performed a case control study for MA by spot urinalysis. The incidence of microalbuminuria was 31.7% of the patients compared with 8.3% of the controls. Of all the traditional risk factors for stroke - age, male gender, hypertension, diabetes, dyslipidemia, smoking showed a positive correlation with the presence of MA (p<0.05). The effect of microalbuminuria and subtypes of ischemic stroke was observed that small vessels stroke (lacunar infract) were more (70%) than large vessels stroke (30%) and effect of microalbuminuria 21.67% higher in small vessels stroke than large vessels stroke. The presence of microalbuminuria appears to independently predict proper clinical outcome following acute stroke.

Trimetazidine in the prevention of contrast induced nephropathy after coronary angiogram.

This prospective randomized, controlled trial was done to evaluate the efficacy of trimetazidine in the prevention of contrast induced nephropathy in patients with raised serum creatinine levels undergoing coronary angiogram. This study was performed in the Department of Cardiology, Bangabandhu Sheikh Mujib Medical University, Dhaka, Bangladesh from April 2009 to March 2010. Contrast-induced nephropathy (CIN) is a serious complication of coronary angiography that is associated with considerably increased mortality and morbidity, including the need for short-term haemodialysis, extended hospitalisation and permanent impairment of renal function. Trimetazidine (TMZ) has been described as a cellular anti-ischaemic agent. This study was trial with 400 patients. Among them 200 patients treated with trimetazidine plus hydration with normal saline and 200 patients (control) given hydration by normal saline only. It was found that the incidence of CIN was significantly (p<0.05) reduced by trimetazidine administration with saline in comparison with saline alone in patients undergoing coronary angiogram (4% vs. 14%).

Non-starch polysaccharide degradation in the gastrointestinal tract of broiler chickens fed commercial-type diets supplemented with either a single dose of xylanase, a double dose of xylanase, or a cocktail of non-starch polysaccharide-degrading enzymes.

The aim of this study was to examine non-starch polysaccharide (NSP) degradation in the gastrointestinal tract of chickens fed a range of commercial-type diets supplemented with a commercial dose of xylanase, a double dose of xylanase or a cocktail of NSP - degrading enzymes. Cobb 500 broilers (n = 1,080) were fed 12 dietary treatments; 4 diets with differing primary grain sources (barley, corn, sorghum, and wheat) and three different enzyme treatments (commercial recommended dose of xylanase (16,000 BXU/kg), a double dose of xylanase (32,000 BXU/kg) or an NSP-degrading enzyme cocktail (xylanase, ß-glucanase, cellulase, pectinase, mannanase, galactanase, and arabinofuranosidase at recommended commercial levels). There were 108 pens, approximately 10 birds per pen, 9 replicates per dietary treatment. The diets were fed as 3 phases, starter (d 0-12), grower (d 12-23), and finisher (d 23-35). On bird age d 12, 23, and 35, performance (total pen body weight, feed intake, and feed conversion ratio corrected for mortality [cFCR]), litter and excreta dry matter content, and ileal and total tract soluble and insoluble NSP degradability and free oligosaccharide digestibility was determined. On d 35, the quantity of NSP in the gizzard, jejunum, ileum and excreta was determined. Results from this study showed that the double xylanase dose and NSP-ase cocktail had positive impacts on starter phase performance in birds fed the corn- and wheat-based diets. In the grower phase in birds fed the barley-based diet, these enzyme treatments improved cFCR and increased litter dry matter content. The NSP-ase cocktail had a negative impact on finisher phase cFCR in birds fed the sorghum-based diet. The double xylanase dose induced a positive impact on NSP degradability and free oligosaccharide digestibility. In conclusion, there appears to be advantages to feeding broilers a double xylanase dose, but lack of consistency when using an NSP-ase cocktail containing many enzymes.

Contribution of individual broilers to variation in amino acid digestibility in soybean meal and the efficacy of an exogenous monocomponent protease.

A total of 72 male Ross 308 broilers were reared to day 34 on a standard wheat and soy-based diet and then offered one of the four semisynthetic experimental diets, comprising two different soybean meal sources either without or with exogenous protease (treatments therefore offered in a 2*2 factorial arrangement). Each experimental diet was fed to 18 individually housed birds from 34 to 37 D after which ileal digesta were collected and digestibility coefficients were calculated. The two soybean meal sources were found to be nutritionally divergent (P < 0.01), with one having the apparent ileal amino acid digestibility coefficient of 0.80 and the other 0.71. Exogenous protease increased (P < 0.01) apparent ileal amino acid digestibility coefficients from 0.74 to 0.77. There were no interactions between soybean meal origin and protease effect. On an individual bird level, there were substantial differences in the capacity to digest amino acids with the mean total amino acid digestibility coefficients from 0.54 to 0.80 for one of the soybean meal samples. Exogenous protease addition reduced the coefficient of variation for total amino acids from 11.4 to 9.1% in one soybean meal and from 7.7 to 6.3% in the other. It can be concluded that soybean meal digestibility varies and that some of this variance is associated with heterogeneity in the digestive capacity of broilers.

Safety and affordability of an elective Saturday list at Pietersburg Hospital, Limpopo, South Africa.

BACKGROUND: The backlog of patients waiting for operations continues to be a problem in many public hospitals in South Africa (SA), with elective surgery procedures being postponed for up to 2 years. OBJECTIVE: To determine the rate of death in hospital or out of hospital within 30 days of an elective procedure performed on a Saturday, and to determine the cost incurred by paying staff members who perform these operations. METHOD: A prospective, observational descriptive cohort study of all patients undergoing inpatient general surgery operations during weekdays and weekends between 1 September 2015 and 31 August 2016 (1 year) at Pietersburg Hospital (PBH), Limpopo, SA. Microsoft Excel 2010 (Microsoft, USA) was used to analyse and derive descriptive statistics. The finance department at the hospital calculated the overtime pay for theatre staff who operated on Saturdays. RESULTS: The study included 1 352 operations (607 elective and 745 emergency procedures). Saturday elective operations contributed 133/607 (22%), and the rate of death for these operations was 1.5%. The most common procedures performed on a Saturday were hernia repair and amputation. The cost for 8 hours of work on a Saturday was ZAR13 900, amounting to a total of ZAR333 600 for 24 Saturdays. CONCLUSION: Performing minor surgery on a Saturday had a mortality rate of 1.5%, and a theatre staff cost of ~ZAR2 317 per patient, excluding surgeons' fees. If surgeons were to be paid the costs would be ZAR3 450 per patient.

Effects of fructose supplementation in chemically defined protein-free medium on development of bovine in vitro fertilized embryos.

The present study evaluated the possible embryotrophic role of fructose supplementation in potassium simplex optimization medium (KSOM) on preimplantation development of bovine in vitro matured and fertilized (IVF) embryos under chemically defined conditions. In Experiment 1, the rates of cleavage (74.0-75.5%) and blastocyst formation (21.0-24.5%) were not affected by the supplementation of fructose in KSOM in absence or presence of glucose. In Experiment 2, the rates of cleavage (71.7-77.3%) and blastocyst formation (19.9-26.3%) did not differ significantly among the concentrations (0.0, 0.2, 1.5, 3.0, 5.6mM) of fructose supplementations in KSOM in presence of glucose. Moreover, the number of total ICM and TE cells, and percentage of ICM to total cell in blastocysts did not differ significantly among the concentrations of fructose supplementations in presence of glucose. In Experiment 3, the rates of cleavage (67.3-74.7%) and blastocyst formation (14.4-19.3%) did not differ significantly among the concentrations (0.0, 0.2, 1.5, 3.0, 5.6mM) of fructose supplementations in KSOM in absence of glucose. Although the number of total and ICM cells, and percentage of ICM to total cells in blastocysts did not differ significantly among the concentrations of fructose supplementations, 1.5mM fructose supplementation in absence of glucose had significantly (P<0.05) higher number of TE cells (106.2) than that of 5.6mM (84.0) supplementation. The study indicates that, fructose up to 5.6mM concentration can be used as an alternative for energy substrate in culture media without any detrimental effect on pre-implantation development in bovine IVF embryos.

Comparative study of endoscopic band ligation and sclerotherapy for treatment of oesophageal varices in cirrhotic patients.

A prospective randomized study was done comparing endoscopic band ligation and sclerotherapy for the treatment of oesophageal varices. Hundred fifty cirrhotic patients with history of bleeding from varices were included in the study. Seventy-five patients got sclerotherapy and 75 got band ligation of varices. Both sclerotherapy and band ligation were continued till obliteration of varices. Clinical as well as the biochemical parameters of the two groups of patients and the severity of the liver diseases were similar in both groups. The mean period of follow up was 337 days for sclerotherapy group and 376 days for band ligation group. The mean time to eradicate the varices was significantly shorter with banding (35+/-4.1 vs 75+/-6.5, p<0.001). The mean number of treatment sessions was significantly less with banding (2.3+/-3.1 vs 5.2+/-2.1, p<.001). Rebleeding was significantly more with sclerotherapy (20 vs. 8 with p<0.01). The variables significantly associated with rebleeding were hypoalbuminaemia (p<.01), and active bleeding at index endoscopy (p<0.001). Different complications like oesophageal ulcer, bleeding from ulcer and the symptoms of oesophageal stricture were more in the sclerotherapy group. There was no significant difference in survival between the two groups. Number of blood transfusions and duration of hospitalization were less in band ligation group. There was no difference in terms of recurrence between the two groups. From the study we suggest that band ligation has less local complications and causes earlier eradication of varices than sclerotherapy. Therefore, band ligation may be the first choice of therapy for oesophageal varices.

Provision of an emergency theatre in tertiary hospitals is cost-effective: Audit and cost of cancelled planned elective general surgical operations at Pietersburg Hospital, Limpopo Province, South Africa.

BACKGROUND: Cancellations of planned elective surgical operations increase financial cost to the patient and the hospital. OBJECTIVES: To determine the rate and reasons for cancellations, estimate the cost incurred by such cancellations and recommend possible solutions. METHODS: We did a prospective descriptive study of cancellations of elective general surgical operations over the 1-year period January - December 2014 in the main theatre at Pietersburg (PTB) Hospital, Limpopo Province, South Africa. All patients listed on the theatre booking slate for elective general surgical operations before the cut-off time of 13h00 on the day before the anticipated operation were included. Epi Infoversion 7 was used to analyse the data and derive the descriptive statistics. RESULTS: There were 537 booked patients (median age 47 years, range 1 - 94); a total of 298 operations were performed, and 239 were cancelled (cancellation rate 44.5%). Reasons for cancellation were as follows: theatre needed for an emergency n=154 (64.4%), theatre equipment failure and lack of consumables n=17 (7.1%), non-theatre equipment failure n=10 (4.2%), prolonged time of operations n=13 (5.4%), abnormal blood results n=8 (3.3%), patient comorbidity and poor general condition n=9 (3.8%), patients absent from the ward n=8 (3.3%), patients not starved n=2 (0.8%), patients' condition improved significantly n=3 (1.3%), nurses' strike n=5 (2.1%), rebooking of cases for senior surgeons or other specialty n=2 (0.8%), and other reasons n=8 (3.3%). The cost per inpatient per day was estimated at ZAR4 890 at PTB Hospital and ZAR2 100 at district hospitals, and the total cost per cancelled operation was ZAR25 860. CONCLUSIONS: Over the 1-year period 44.5% of elective operations at PTB Hospital were cancelled, 64.4% because the theatre was needed for an emergency operation. We recommend that a theatre dedicated to emergencies be opened at PTB Hospital. The cost incurred due to cancellations was about ZAR6 million for the hospital, with additional cost and emotional trauma for the patients.

An approach for producing transgenic cloned cows by nuclear transfer of cells transfected with human alpha 1-antitrypsin gene.

In an attempt to produce transgenic cloned cows secreting alpha 1-antitrypsin (alpha1-AT) protein into milk, bovine cumulus cells were transfected with a plasmid containing an alpha1-AT gene and green fluorescent protein (GFP) reporter gene using Fugene 6 as a lipid carrier. The GFP-expressing cells were selected and transferred into enucleated bovine oocytes. Couplets were fused, chemically activated and cultured. Developmental competence was monitored and the number of inner cell mass (ICM) and trophectoderm (TE) cells in blastocysts were counted after differential staining. The percentage of blastocysts was lower (P < 0.05) in transgenic cloned embryos compared to non-transgenic cloned embryos (23% versus 35%). No difference in the numbers of ICM and TE cells between the two groups of embryos was observed. One or two GFP-expressing blastocysts were transferred into the uterus of each recipient cow. Out of 49 recipient cows, three pregnancies were detected by non-return estrus and rectal palpation. However, the pregnancies failed to maintain to term; two fetuses were aborted at Day 60 and 150, respectively, and one fetus at Day 240. The genomic DNA from the aborted fetus was amplified by polymerase chain reaction (PCR) to investigate integration of the transgene in the fetus. The expected PCR product was sequenced and was identical to the sequence of alpha1-AT transgene. In conclusion, the present study demonstrated that developmental competence of cloned embryos derived from transgenic donor cells was lower than embryos derived from non-transfected donor cells. Although we failed to obtain a viable transgenic cloned calf, integration of alpha1-AT gene into the fetus presents the possibility of producing transgenic cloned cows by somatic cell nuclear transfer.

Management of sigmoid volvulus in Polokwane-Mankweng Hospital.

OBJECTIVE: To evaluate the outcome of treatment of patients with sigmoid volvulus in the Polokwane-Mankweng Hospital and to identify the best management options for these patients. METHODS: A retrospective study was undertaken of 85 patients with sigmoid volvulus treated in Polokwane-Mankweng Hospital during the period July 1997-May 2004. RESULTS: In total, 85 patients were evaluated (77 males and 8 females, male/female ratio 9:1). The age range was 7-80 years (mean 42 years). Sigmoidoscopic derotation was attempted in 17 patients, and was successful in 10 patients. Laparotomy was done in 84 patients, viz. 75 emergencies and 9 electives. During laparotomy, gangrenous sigmoid colon was found in 30 patients and viable sigmoid in 54 patients. Resection with primary anastomosis was done in 44 patients. Hartmann's procedure was performed in 33 patients. Sigmoidopexy was done in 7 patients. Total hospital mortality was 6% (5 deaths). Mortality in the 84 operated cases was 5% (4 deaths). CONCLUSIONS: There was no mortality in patients undergoing elective resection and primary anastomosis after successful preoperative deflation and in patients with viable sigmoid volvulus who underwent an emergency Hartmann's procedure. There was low mortality in those patients with resection and primary anastomosis on viable sigmoid (3%, 1:39). The highest mortality (1:5) occurred in cases of resection and primary anastomosis of gangrenous sigmoid colon.

Nodular thyroid disease and thyroid malignancy: Experience at Polokwane Mankweng Hospital Complex, Limpopo Province, South Africa.

BACKGROUND: Nodular thyroid disease is common throughout the world. Numbers of patients with goitre are increasing worldwide, as also noted in Limpopo Province, South Africa (SA). Globally, thyroid nodules have been reported in 4-7% of the population on neck palpation and in 30-50% by ultrasonography. OBJECTIVES: To review the profile of thyroid disease in patients with goitre presenting to the Department of Surgery at Polokwane Mankweng Hospital Complex (PMHC), Limpopo, SA, to characterise the pattern of malignancy in patients with goitre, and to determine the most common thyroid cancer. METHOD: A 6-year retrospective study (2003-2008) of all patients with thyroid nodules who underwent thyroid surgery at PMHC. RESULTS: The study group included 90 patients (mean age 45 years, range 4-80). The male-to-female ratio was 1:17 (5 men, 85 women). Of these patients, 80 (89.9%) had benign lesions, of which 52 (57.8% of the total) were adenomas, 25 (27.8%) multinodular goitres (MNGs), 2 (2.2%) hyperplastic nodules and 1 (1.1%) Hashimoto's thyroiditis. Ten patients (11.1%) had malignant lesions (7 follicular carcinomas and 3 papillary carcinomas), of which 2 were found in MNGs. CONCLUSIONS: Adenoma and MNG were the predominant non-malignant conditions (85.6%). The prevalence of thyroid cancer in our study was 11.1%, and of all 90 patients, 7.8% had follicular carcinoma. The risk of malignancy in MNG was 8.9%. Rates of thyroid nodules and carcinoma were highest in women aged 41-60 years. We advocate that total thyroidectomy be considered for MNG, because MNG can harbour incidental carcinoma.

Expression of menin in parathyroid tumors.

The multiple endocrine neoplasia type 1 (MEN1) gene seems to be a tumor suppressor that encodes a 610-amino acid protein termed menin and that plays an important role in the development of MEN1 syndrome. Recent reports indicate that heterozygous germline mutations of this gene are responsible for the disease onset of MEN1. In this study we examined the expression of menin in parathyroid tumors from primary hyperparathyroidism (PHP), secondary hyperparathyroidism (SHP), and MEN1 and thyroid tumors including Basedow's disease, thyroid cancer, and adrenocortical tumors. Both ribonucleic acid and protein from these tumors were applied to RT-PCR and Western blotting, respectively. Primers for RT-PCR were designed to amplify the sequence between exons 2 and 3 of the MEN1 gene. Specific antibody against menin was generated in guinea pigs immunized with the recombinant peptide from amino acid residues 443-535 of menin made by using glutathione-S-transferase (GST) gene fusion. Menin messenger ribonucleic acid was strongly expressed on RT-PCR analysis in the parathyroid tumors from both PHP and SHP. Western blotting revealed a specific band of approximately 67 kDa in parathyroid tumors from PHP and SHP, with a much weaker such band detected in thyroid tumors. Menin expression was down-regulated in MEN1 samples, including nonsense mutation and deletion mutant. These findings suggest that menin is predominantly synthesized and stored in parathyroid tumors resulting from PHP and SHP.

Preimplantational embryo development and incidence of blastomere apoptosis in bovine somatic cell nuclear transfer embryos reconstructed with long-term cultured donor cells.

This study was performed to investigate whether types and/or age of donor cells affect preimplantational embryo development and the incidence of apoptosis in bovine somatic cell nuclear transfer (SCNT) embryos. Bovine fetal or adult ear fibroblasts were isolated, cultured in vitro and categorized into fresh or long-term cultured cells in terms of population doublings (PD): in fetal fibroblasts, <16 being considered fresh and >50 being long-term cultured; in adult ear fibroblasts, <16 being considered fresh and >30 being long-term cultured. Bovine oocytes from slaughterhouse ovaries were matured in TCM-199, enucleated and reconstructed by SCNT. The reconstructed oocytes were fused, chemically activated, and cultured in modified synthetic oviduct fluid (mSOF) at 39 degrees C in a humidified atmosphere of 5% CO(2) air for 7 days. The early development of SCNT embryos was monitored under a microscope and the quality of blastocysts was assessed by differential counting of inner cell mass (ICM) and trophectoderm (TE) cells and by apoptosis detection in blastomeres using a terminal deoxynucleotidyl transferase-mediated d-UTP nick end-labeling (TUNEL) assay. As results, types and/or age of donor cells did not affect the rate of blastocyst formation and the number of ICM and TE cells. However, a significant increase in apoptotic blastomeres was observed in SCNT embryos reconstructed with long-term cultured fetal or adult ear fibroblasts compared to those in SCNT embryos derived from fresh fetal or adult ear fibroblasts. In conclusion, these results indicated that the long-term culture of donor cells caused increased the incidence of apoptosis in bovine SCNT embryos but did not affect the developmental competence and the cell number of blastocysts.

Effect of protein supplementation in potassium simplex optimization medium on preimplantation development of bovine non-transgenic and transgenic cloned embryos.

The present study evaluated the effect of protein supplementation in potassium simplex optimization medium (KSOM) on bovine preimplantation embryo development. The in vitro fertilized (IVF) (Experiment 1), non-transgenic (Experiment 2) and transgenic cloned embryos (Experiment 3) were cultured for 192 h in KSOM supplemented with 0.8% BSA (KSOM-BSA), 10% FBS (KSOM-FBS) or 0.01% PVA (KSOM-PVA). Transfected cumulus cells with an expression plasmid for human alpha1-antitrypsin gene and a green fluorescent protein (GFP) marker were used to produce transgenic cloned embryos. Modified synthetic oviductal fluid (mSOF) supplemented with 0.8% BSA (mSOF-BSA) was used as a control medium. In Experiment 1, cleavage rate was significantly (P < 0.05) lower (69.1%) in IVF embryos cultured in KSOM-FBS than in KSOM-BSA (80.3%). The rate of hatching/hatched blastocyst formation was significantly (P < 0.05) lower in embryos cultured in KSOM-PVA than in KSOM-FBS (2.2% versus 10.8%). Blastocysts cultured in KSOM-FBS contained significantly (P < 0.06) higher numbers of inner cell mass cells (50.4 +/- 20.2) than those cultured in mSOF-BSA (36.9 +/- 19.2). In Experiment 2, the rate of blastocyst formation was significantly (P < 0.05) lower (20.5%) in embryos cultured in KSOM-PVA than in other culture media (33.3-38.5%). The rate of hatching/hatched blastocysts was significantly (P < 0.05) lower in KSOM-PVA (13.9%) and KSOM-FBS (17.1%) than in KSOM-BSA (30.8%) and mSOF-BSA (33.9%). The numbers of total and trophectoderm cells (104.6 +/- 32.2 and 71.7 +/- 25.5, respectively) were significantly (P < 0.05) lower in blastocysts cultured in KSOM-PVA than in KSOM-BSA (125.7 +/- 39.7 and 91.7 +/- 36.2, respectively). In Experiment 3, no significant differences in embryo development, GFP expression and blastocyst cell numbers were observed among the culture groups. In conclusion, the present study demonstrated that KSOM and mSOF supplemented with BSA were equally effective in supporting development of bovine non-transgenic and transgenic cloned embryos. Moreover, different developmental competence in response to protein supplementation of KSOM was observed between bovine non-transgenic and transgenic cloned embryos.

Effect of transfection and passage number of ear fibroblasts on in vitro development of bovine transgenic nuclear transfer embryos.

The objective of this study was to determine if the transfection of human prourokinase (ProU) gene and passage number of transfected ear fibroblasts affected in vitro development of bovine transgenic nuclear transfer (NT) embryos. An expression plasmid for human ProU was constructed by inserting a bovine beta-casein promoter, a green fluorescent protein (GFP) marker and human ProU gene into a pcDNA3 plasmid and transfected into bovine ear fibroblasts using a lipid mediated method. Abattoir derived oocytes were enucleated at 18-20 hr post maturation and a single donor cell was transferred into the perivitelline space of a recipient oocyte. After fusion and activation, the couplets were cultured in modified synthetic oviductal fluid (mSOF) medium for 168 hr. In Experiment 1, significantly lower rate in blastocysts formation (10.3%) was observed in transfected donor cells at early passage than that in nontransfected counterparts (22.1%, P<0.05). In Experiment 2, development to blastocysts and GFP expression in blastocysts were not significantly different between early (3-7) and late (8-12) passage donor cells (10.3 vs. 11.3% and 54.5 vs. 41.7%, respectively). This study indicates that in vitro development of bovine transgenic NT embryos is negatively influenced by transfection of human ProU gene into donor fibroblasts. However, passage number of transfected ear fibroblasts does not affect in vitro development of bovine transgenic NT embryos.