Bio-scaffolds in organ-regeneration: Clinical potential and current challenges.

Cadaveric organ transplantation represents the definitive treatment option for end-stage disease but is restricted by the shortage of clinically-viable donor organs. This limitation has, in part, driven current research efforts for in vitro generation of transplantable tissue surrogates. Recent advances in organ reconstruction have been facilitated by the re-purposing of decellularized whole organs to serve as three-dimensional bio-scaffolds. Notably, studies in rodents indicate that such scaffolds retain native extracellular matrix components that provide appropriate biochemical, mechanical and physical stimuli for successful tissue/organ reconstruction. As such, they support the migration, adhesion and differentiation of reseeded primary and/or pluripotent cell populations, which mature and achieve functionality through short-term conditioning within specialized tissue bioreactors. Whilst these findings are encouraging, significant challenges remain to up-scale the present technology to accommodate human-sized organs and thereby further the translation of this approach towards clinical use. Of note, the diverse structural and cellular composition of large mammalian organ systems mean that a "one-size fits all" approach cannot be adopted either to the methods used for their decellularization or the cells required for subsequent re-population, to create fully functional entities. The present review seeks to highlight the clinical potential of decellularized organ bio-scaffolds as a route to further advance the field of tissue- and organ-regeneration, and to discuss the challenges which are yet to be addressed if such a technology is ever to become a credible rival to conventional organ allo-transplantation.

Superior mesenteric artery syndrome: a single centre experience of laparoscopic duodenojejunostomy as the operation of choice.

INTRODUCTION The superior mesenteric artery (SMA) syndrome, or Wilkie's syndrome, is a rare cause of postprandial epigastric pain, vomiting and weight loss caused by compression of the third part of the duodenum as it passes beneath the proximal superior mesenteric artery. The syndrome may be precipitated by sudden weight loss secondary to other pathologies, such as trauma, malignancy or eating disorders. Diagnosis is confirmed by angiography, which reveals a reduced aorto-SMA angle and distance, and contrast studies showing duodenal obstruction. Conservative management aims to increase intra-abdominal fat by dietary manipulation and thereby increase the angle between the SMA and aorta. Where surgery is indicated, division of the ligament of Treitz, anterior transposition of the third part of the duodenum and duodenojejunostomy have been described. METHODS We present four cases of SMA syndrome where the intention of treatment was laparoscopic duodenojejunostomy. The procedure was completed successfully in three patients, who recovered quickly with no short-term complications. A fourth patient underwent open gastrojejunostomy (complicated by an anastomotic bleed) when dense adhesions prevented duodenojejunostomy. CONCLUSIONS The superior mesenteric artery syndrome should be considered in patients with epigastric pain, prolonged vomiting and weight loss. Laparoscopic duodenojejunostomy is a safe and effective operation for management of the syndrome. A multi-speciality team approach including gastrointestinal, vascular and radiological specialists should be invoked in the management of these patients.

Open Abdominal Aortic Aneurysm Replacement in the Awake Patient.

Nonintubated aortic surgery using various techniques has been reported, but despite publication of favorable outcomes in select patient groups, awake aortic surgery remains unpopular. Our patient had an abdominal aortic aneurysm that was unsuitable for endovascular repair. Because of the significant respiratory disease, general anesthesia represented an unacceptably high risk. As a result, he underwent open AAA repair via a retroperitoneal approach with the aid of epidural anesthesia. Here, we highlight the benefits of the procedure which offer a select cohort of patients the chance of life-saving surgery.

Field study of dried blood spot specimens for HIV-1 drug resistance genotyping.

Dried blood spots (DBS) are an alternative specimen type for HIV drug resistance genotyping in resource-limited settings. Data relating to the impact of DBS storage and shipment conditions on genotyping efficiency under field conditions are limited. We compared the genotyping efficiencies and resistance profiles of DBS stored and shipped at different temperatures to those of plasma specimens collected in parallel from patients receiving antiretroviral therapy in Uganda. Plasma and four DBS cards from anti-coagulated venous blood and a fifth card from finger-prick blood were prepared from 103 HIV patients with a median viral load (VL) of 57,062 copies/ml (range, 1,081 to 2,964,191). DBS were stored at ambient temperature for 2 or 4 weeks or frozen at -80 °C and shipped from Uganda to the United States at ambient temperature or frozen on dry ice for genotyping using a broadly sensitive in-house method. Plasma (97.1%) and DBS (98.1%) stored and shipped frozen had similar genotyping efficiencies. DBS stored frozen (97.1%) or at ambient temperature for 2 weeks (93.2%) and shipped at ambient temperature also had similar genotyping efficiencies. Genotyping efficiency was reduced for DBS stored at ambient temperature for 4 weeks (89.3%, P = 0.03) or prepared from finger-prick blood and stored at ambient temperature for 2 weeks (77.7%, P < 0.001) compared to DBS prepared from venous blood and handled similarly. Resistance profiles were similar between plasma and DBS specimens. This report delineates the optimal DBS collection, storage, and shipping conditions and opens a new avenue for cost-saving ambient-temperature DBS specimen shipments for HIV drug resistance (HIVDR) surveillances in resource-limited settings.

Enhanced reaction rates in NDP analysis with neutron scattering.

Neutron depth profiling (NDP) makes accessible quantitative information on a few isotopic concentration profiles ranging from the surface into the sample a few micrometers. Because the candidate analytes for NDP are few, there is little interference encountered. Furthermore, neutrons have no charge so mixed chemical states in the sample are of no direct concern. There are a few nuclides that exhibit large probabilities for neutron scattering. The effect of neutron scattering on NDP measurements has not previously been evaluated as a basis for either enhancing the reaction rates or as a source of measurement error. Hydrogen is a common element exhibiting large neutron scattering probability found in or around sample volumes being analyzed by NDP. A systematic study was conducted to determine the degree of signal change when neutron scattering occurs during analysis. The relative signal perturbation was evaluated for materials of varied neutron scattering probability, concentration, total mass, and geometry. Signal enhancements up to 50% are observed when the hydrogen density is high and in close proximity to the region of analysis with neutron beams of sub thermal energies. Greater signal enhancements for the same neutron number density are reported for thermal neutron beams. Even adhesive tape used to position the sample produces a measureable signal enhancement. Because of the shallow volume, negligible distortion of the NDP measured profile shape is encountered from neutron scattering.

Outbreak of type A foodborne botulism at a boarding school, Uganda, 2008.

Botulism has rarely been reported in Africa. In October 2008, botulism was reported in three Ugandan boarding-school students. All were hospitalized and one died. A cohort study was performed to assess food exposures among students, and clinical specimens and available food samples were tested for botulinum toxin. Three case-patients were identified; a homemade, oil-based condiment was eaten by all three. In the cohort study, no foods were significantly associated with illness. Botulinum toxin type A was confirmed in clinical samples. This is the first confirmed outbreak of foodborne botulism in Uganda. A homemade, oil-based condiment was the probable source. Consumption of homemade oil-based condiments is widespread in Ugandan schools, putting children at risk. Clinicians and public health authorities in Uganda should consider botulism when clusters of acute flaccid paralysis are seen. Additionally, schools should be warned of the hazard of homemade oil-based condiments, and take steps to prevent their use.

Webuye Health and Demographic Surveillance Systems Baseline Survey of Soil-Transmitted Helminths and Intestinal Protozoa among Children up to Five Years.

Background. The intestinal parasitic infections (IPIs) are globally endemic, and they constitute the greatest cause of illness and disease worldwide. Transmission of IPIs occurs as a result of inadequate sanitation, inaccessibility to potable water, and poor living conditions. Objectives. To determine a baseline prevalence of IPIs among children of five years and below at Webuye Health and Demographic Surveillance (HDSS) area in western Kenya. Methods. Cross-sectional survey was used to collect data. Direct saline and formal-ether-sedimentation techniques were used to process the specimens. Descriptive and inferential statistics such as Chi-square statistics were used to analyze the data. Results. A prevalence of 52.3% (417/797) was obtained with the male child slightly more infected than the female (53.5% versus 51%), but this was not significant (χ (2) = 0.482, P > 0.05). Giardia lamblia and Entamoeba histolytica were the most common pathogenic IPIs with a prevalence of 26.1% (208/797) and 11.2% (89/797), respectively. Soil-transmitted helminths (STHs) were less common with a prevalence of 4.8% (38/797), 3.8% (30/797), and 0.13% (1/797) for Ascaris lumbricoides, hookworms, and Trichuris trichiura, respectively. Conclusions. Giardia lamblia and E. histolytica were the most prevalent pathogenic intestinal protozoa, while STHs were less common. Community-based health promotion techniques are recommended for controlling these parasites.

Rotational co-culture of clonal ß-cells with endothelial cells: effect of PPAR-γ agonism in vitro on insulin and VEGF secretion.

AIM: Delayed graft revascularization impedes the success of human islet transplantation. This study utilized rotational co-culture of insulin secreting ß-cells with human umbilical vein endothelial cells (HUVECs) and a peroxisome proliferator-activated receptor gamma (PPAR-γ) agonist to promote insulin and vascular endothelial growth factor (VEGF) secretory function. METHODS: Clonal BRIN-BD11 (D11) cells were maintained in static culture (SC) and rotational culture (RC) ± HUVEC and ± the TZD (thiazolidinedione) rosiglitazone (10 mmol/l) as a specific PPAR-γ agonist. HUVECs were cultured in SC and RC ± D11 and ± TZD. D11 insulin secretion was induced by static incubation with low glucose (1.67 mmol/l), high glucose (16.7 mmol/l) and high glucose with 10 mmol/l theophylline (G+T) and assessed by enzyme-linked immunosorbent assay (ELISA). HUVEC proliferation was determined by ATP luminescence, whereas VEGF secretion was quantified by ELISA. Co-cultured cells were characterized by immunostaining for insulin and CD31. RESULTS: D11 SC and RC showed enhanced insulin secretion in response to 16.7 mmol/l and G+T (p < 0.01); without significant alteration by the TZD. Co-culture with HUVEC in SC and RC also increased D11 insulin secretion when challenged with 16.7 mmol/l and G+T (p < 0.01), and this was slightly enhanced by the TZD. The presence of HUVEC increased D11 SC and RC insulin secretion in response to high glucose and G+T, respectively (p < 0.01). Addition of the TZD increased SC and RC HUVEC ATP content (p < 0.01) and VEGF production (p < 0.01) in the presence and absence of D11 cells. CONCLUSIONS: Rotational co-culture of insulin secreting cells with endothelial cells, and exposure to a PPAR-γ agonist may improve the prospects for graft revascularization and function after implantation.

Sustained insulin secretory response in human islets co-cultured with pancreatic duct-derived epithelial cells within a rotational cell culture system.

AIMS/HYPOTHESIS: Loss of the trophic support provided by surrounding non-endocrine pancreatic cell populations underlies the decline in beta cell mass and insulin secretory function observed in human islets following isolation and culture. This study sought to determine whether restoration of regulatory influences mediated by ductal epithelial cells promotes sustained beta cell function in vitro. METHODS: Human islets were isolated according to existing protocols. Ductal epithelial cells were harvested from the exocrine tissue remaining after islet isolation, expanded in monolayer culture and characterised using fluorescence immunocytochemistry. The two cell types were co-cultured under conventional static culture conditions or within a rotational cell culture system. The effect of co-culture on islet structural integrity, beta cell mass and insulin secretory capacity was observed for 10 days following isolation. RESULTS: Human islets maintained under conventional culture conditions exhibited a characteristic loss in structural integrity and functional viability as indicated by a diminution of glucose responsiveness. By contrast, co-culture of islets with ductal epithelial cells led to preserved islet morphology and sustained beta cell function, most evident in co-cultures held within the rotational cell culture system, which showed a significantly (p < 0.05) greater insulin secretory response to elevated glucose compared with control islets. Similarly, insulin/protein ratio data suggested that the presence of ductal epithelial cells is beneficial for the maintenance of beta cell mass. CONCLUSIONS/INTERPRETATION: The data indicate a supportive role for ductal epithelial cells in islet viability. Further characterisation of the regulatory influences may lead to novel strategies to improve long-term beta cell function both in vitro and following islet transplantation.

Aetiology of diarrhoea among persons with HIV and their family members in rural Uganda: a community-based study.

OBJECTIVES: To identify pathogens associated with diarrhoea in HIV-infected persons and their HIV-uninfected family members. DESIGN: Prospective cohort study. SETTING: Rural community in eastern Uganda. SUBJECTS: Eight hundred and seventy nine HIV-infected adults (74% females and median age 35 years (IQR, 29-41) and 2771 HIV-uninfected family members (51% females and median age 11 years (IQR 6-16) were included. MAIN OUTCOME MEASURES: Using microscopy and culture, stools were tested for parasites, bacteria and bacterial-antimicrobial-susceptibility. Logistic regression models, adjusting for age, CD4 cells, season, household clustering and use of safe-water system were used for relationships between pathogens, diarrhoea and HIV. RESULTS: Persons with HIV had similar pathogens in diarrhoeal (69%) and nondiarrhoeal stools (57%). Most diarrhoea was not associated with identifiable aetiology; the population attributable risk of diarrhoea for known diarrhoea pathogens was 32%. Enteric bacteria (19%), enteropathogenic or enterotoxigenic E. coli (8%), Aeromonas species (7%), Strongyloides stercoralis (8%) and Cryptosporidium parvum (5%). HIV-infected, stools had more Cryptosporidium parvum than HIV-uninfected (OR 2.64, 95% CI 1.43-4.87). Most bacteria were resistant to commonly used antimicrobials irrespective of HIV status. CONCLUSIONS: Irrespective of HIV-status, aetiologies of majority of their diarrhoea in Uganda cannot be identified by microscopy and culture. Bacterial pathogens isolated have high resistance to common antimicrobials. Empiric treatment should be tailored to local bacterial-resistance patterns.

Role of the laboratory in ensuring global access to ARV treatment for HIV-infected children: consensus statement on the performance of laboratory assays for early infant diagnosis.

A two day meeting hosted by the World Health Organization (WHO) and the U.S. Centers for Disease Control and Prevention (CDC) was held in May 2006 in Entebbe, Uganda to review the laboratory performance of virologic molecular methods, particularly the Roche Amplicor DNA PCR version 1.5 assay, in the diagnosis of HIV-1 infection in infants. The meeting was attended by approximately 60 participants from 17 countries. Data on the performance and limitations of the HIV-1 DNA PCR assay from 9 African countries with high-burdens of HIV/AIDS were shared with respect to different settings and HIV- subtypes. A consensus statement on the use of the assay for early infant diagnosis was developed and areas of needed operational research were identified. In addition, consensus was reached on the usefulness of dried blood spot (DBS) specimens in childhood as a means for ensuring greater accessibility to serologic and virologic HIV testing for the paediatric population.

Presumed interaction of fusidic acid with simvastatin.

A 63-year-old man was admitted 6 weeks after an elective abdominal aortic aneurysm repair following which methicillin resistant Staphylococcus aureus (MRSA) had been cultured from the aneurysmal sac. He had been commenced on a course of fusidic acid at discharge in addition to his ongoing statin prescription and presented 4 weeks later with symptoms consistent with rhabdomyolysis. Severe rhabdomyolysis was confirmed and despite prolonged and complicated critical care management, his treatment was unsuccessful. Extensive investigations ruled out other known causes of this clinical presentation and failed to identify any other precipitating cause of rhabdomyolysis. We believe the most likely cause was hepatic inhibition of the CYP3A4 hepatic isoenzyme by fusidic acid resulting in an acute severe rise in plasma simvastatin level and extensive myocellular damage. Increasing MRSA colonisation and infection rates together with increased statin usage have the potential to increase the incidence of this presumed drug interaction.

Prevalence and screening costs of hepatitis C virus among Ugandan blood donors.

BACKGROUND: Screening donated blood for hepatitis C virus (HCV) is important for HCV prevention and is routinely practiced in North America and Europe. However, in many African countries little is known about HCV prevalence or cost-effectiveness of HCV antibody (anti-HCV) screening. METHODS: We investigated 2592 plasma specimens collected consecutively from blood donors in central Uganda in 1999. Routine screening by the blood bank included human immunodeficiency virus (HIV), hepatitis B surface antigen (HBsAg), and syphilis. To assess HCV prevalence and cost-effectiveness of testing, specimens were additionally tested for anti-HCV IgG by enzyme immunosorbent assay (EIA). Specimens repeatedly reactive (RR) on EIA were tested with a recombinant immunoblot assay (RIBA). RESULTS: Overall, 107 (4.1%) specimens were HCV EIA RR. Fifteen EIA RR specimens (0.6%, 95% confidence interval = 0.3-0.9%) were RIBA positive and 47 (1.8%) were RIBA indeterminate. Most (80%) RIBA-positive specimens were non-reactive for HIV, HBsAg, and syphilis. RIBA positivity was not associated with donor age, sex, number of donations, HIV, or HBsAg positivity. Costs of screening donors for anti-HCV by using EIA were estimated at US Dollars 782 per potential transfusion-associated HCV infection (exposure to RIBA-positive blood) averted. CONCLUSIONS: Current screening tests for other infections are ineffective in removing HCV-positive donations. Testing costs are considerable; cost-effectiveness of identifying HCV-infected donors will be critical in decision making about HCV screening in Uganda.

Prognostic significance of raised cardiac troponin T in patients presenting with acute limb ischaemia.

OBJECTIVE: To study the relation between serum cardiac troponin T (cTnT) and mortality in patients presenting with acute limb ischaemia secondary to an embolism. MATERIAL AND METHODS: A two years prospective study of all patients admitted to the vascular unit with a diagnosis of acute limb ischaemia secondary to an embolism. On admission all patients had an ECG. A blood sample was taken for measurement of cTnT, CRP, serum biochemistry, full blood count and clotting. All embolectomies were performed under local anaesthesia. Patients were followed until discharge from hospital and up to twelve months after surgery. RESULTS: There were 37 patients with lower limb and 2 patients with upper limb ischaemia. Twenty four patients were female and fifteen were male, with the mean age of 76 years (50-95) for women and 84 years (77-90) for men. Seventeen patients (44%) had a raised cTnT. The patients with raised cTnT were older than those with normal cTnT [86y (77-92) vs 77y (51-95), p = 0.01, t test]. The mean cTnT was 0.20 microg/L (range: 0.11-0.27). Only two patients with raised cTnT gave a history of chest pains. All of the patients with an elevated cTnT had also raised CRP. There was no significant difference in the serum creatinine in the group of patients with elevated cTnT compared to those with normal cTnT [112 micromol/L (range 98-159) vs 119 micromol/L (range: 47-177), p = ns]. The cumulative survival for cTnT+ patients at 7 days was 53% and that of cTnT- patients was 100%. The cumulative survival for cTnT+ and cTnT- patients was statistically different (p = 0.0000, chi2 = 13.1, Log Rank test). Using regression analysis, an elevated cTnT was found to be an independent predictor of outcome. CONCLUSION: A significant proportion of patients presenting with an acutely ischaemic limb have an elevated cTnT. An elevated cTnT may be an early marker of overall disease severity and a predictor of outcome.

Corneo-conjunctival carcinoma in Uganda.

AIMS: Since 1990, the incidence of conjunctival neoplasia has more than tripled in Uganda. It is known to be associated with exposure to solar ultraviolet radiation and to infection with the human immunodeficiency virus-1 (HIV). However, little is known about the most effective treatments. In this study, we report surgical outcomes among people with corneo-conjunctival squamous neoplasia in Uganda and investigate the role of HIV infection and other factors in the aetiology of the tumour. METHODS: Country-wide enrolment of participants; removal and histology of suspect lesions; HIV counselling and testing; home visiting of participants to determine outcomes. RESULTS: In 67 months between 1995 and 2001, 476 participants were enrolled (262 female, 214 male, median age 32 years). A total of 463 (97%) had eye-conserving removal of the lesion and 13 had other surgery. For 414, the histology was squamous neoplasia (184 invasive carcinoma, 230 intraepithelial). The prevalence of HIV infection in cases was 64%. In all, 96% were followed up for a median period of 32 months (range 0-81) after eye-conserving surgery during which time 13 (3.2%) had a recurrence. CONCLUSIONS: Surgery resulted in a low recurrence rate during the follow-up period and had minimal complications. The prevalence of HIV among cases was higher than expected on the basis of data from the general population, although about a third of cases were HIV-negative and had normal CD4 counts. No new cofactors were identified.

Evaluating the role of human papillomaviruses in conjunctival neoplasia.

Mucosal, cutaneous and Epidermodysplasia verruciformis (EV)-related human papillomaviruses (HPVs) were searched by broad-spectrum PCR in 86 conjunctival neoplasia biopsies and 63 conjunctival non-neoplastic control tissue from Ugandan subjects. Seven different EV-related HPV types, including a putative new HPV, and two mucosal HPVs were detected in 25% (14 out of 56) of HIV-positive, in 10% (three out of 30) of HIV-negative conjunctival neoplasia samples, and rarely (0-1.6%) in control subjects. The absence of high-risk HPVs and the low detection frequency of EV-related HPV types in more advanced tumour stages (10%) raise doubts about their role in conjunctival carcinomas.

Chronic herpes simplex virus type-2 infection and HIV viral load.

By December 2003, the estimated adult HIV/AIDS prevalence rate in sub-Saharan Africa was 7.5-8.5%, and rates of herpes simplex virus type-2 (HSV-2) infection among adults aged >30 years ranged from 60% to 82%. However, little is known about the natural history of HIV/HSV-2 co-infection in this population. We evaluated HIV viral load and CD4+ cell counts among persons with and without chronic HSV-2 co-infection in a cross-sectional study of HIV-infected persons not receiving antiretroviral therapy. HSV-2 and HIV co-infection was associated with a 0.3 log copies/mL higher HIV viral load compared with persons without HSV-2 infection (P=0.014). Chronic HSV-2 infection may have a negative effect on the clinical course of persons with HIV.

Are organ preservation solutions useful for the storage of isolated human islets?

Clinical islet transplantation invariably requires more than one donor per recipient. Delay between transplants could be reduced if islets were stored and transported between centers. This study assessed viability and response to glucose of isolated human islets after storage in tissue culture medium (TCM) 199 at 30 degrees C (control), TCM 199 at 22 degrees C (RT), University of Wisconsin solution @ 4 degrees C (UW), or Eurocollins Solution at 4 degrees C (EC) and compared 18 hours storage (group 1) or overnight culture followed by 4 hours storage (group 2). Insulin stimulation index (SI) (mean +/- SD, n = 5), after 1 hour glucose static challenge was not significantly different (P >.05) from islets in group 1 stored in RT 1.76 +/- 1.08 or EC 1.14 +/- 0.29 versus control 2.41 +/- 1.13 or group 2, RT 1.73 +/- 0.51, EC 2.07 +/- 0.63 versus control 2.12 +/- 0.58. However, SI UW was significantly lower (P <.05) than the control in group 1 (1.19 +/- 0.30) and group 2 (1.36 +/- 0.34). Islet viability represented by the ATP/ADP ratio (mean +/- SD, n = 5) was not significantly different after storage in RT 0.201 +/- 0.159; EC 0.205 +/- 0.123; or UW 0.611 +/- 0.992 versus the control 0.223 +/- 0.158 in group 1, and RT 0.178 +/- 0.055; EC 0.137 +/- 0.018; or UW 0.173 +/- 0.085, compared with the control 0.199 +/- 0.069 in group 2. We conclude, organ preservation solutions EC and UW do not have an advantage over TCM 199 for the storage of isolated human islets.