Maureen Duffield

Characterization of a CD46 transgenic pig and protection of transgenic kidneys against hyperacute rejection in non-immunosuppressed baboons

Abstract:  Human membrane cofactor protein (CD46) controls complement activation and when expressed sufficiently as a transgene protects xenografts against complement‐mediated rejection, as shown here using non‐immunosuppressed baboons and heterotopic CD46 transgenic pig kidney xenografts. This report is of a carefully engineered transgene that enables high‐level CD46 expression. A novel CD46 minigene was validated by transfection and production of a transgenic pig line. Pig lymphocytes were tested for resistance to antibody and complement‐mediated lysis, transgenic tissues were characterized for CD46 expression, and kidneys were transplanted to baboons without immunosuppression. Absorption of anti‐Galα(1,3)Gal epitope (anti‐GAL) serum antibodies was measured. Transgenic pigs expressed high levels of CD46 in all tissues, especially vascular endothelium, with stable expression through three generations that was readily monitored by flow cytometry of transgenic peripheral blood mononuclear cells (PBMC). Transgenic PBMC pre‐sensitized with antibody were highly resistant to human complement‐mediated lysis which readily lysed normal pig PBMC. Normal pig kidneys transplanted without cold ischemia into non‐immunosuppressed adult baboons survived a median of 3.5 h (n = 7) whereas transgenic grafts (n = 9), harvested at ∼24‐h intervals, were either macroscopically normal (at 29, 48 and 68 h) or showed limited macroscopic damage (median > 50 h). Microscopic assessment of transplanted transgenic kidneys showed only focal tubular infarcts with viable renal tissue elsewhere, no endothelial swelling or polymorph adherence and infiltration by lymphocytes beginning at 3 days. Coagulopathy was not a feature of the histology in four kidneys not rejected and assessed at 48 h or later after transplantation. Baboon anti‐GAL serum antibody titers were high before transplantation and, in one extensively analyzed recipient, reduced ∼8‐fold within 5.5 h. The data demonstrate that a single CD46 transgene controls hyperacute kidney graft rejection in untreated baboons despite the presence of antibody and complement deposition. The expression levels, tissue distribution and in vitro functional tests indicate highly efficient CD46 function, controlling both classical and alternative pathway complement activation, which suggests it might be the complement regulator of choice to protect xenografts.

The spectrum of renal histologies seen in HIV with outcomes, prognostic indicators and clinical correlations

BACKGROUND Two hundred and twenty-one HIV-positive renal biopsies were analysed from Groote Schuur Hospital to determine outcomes and prognostic indicators based on histology and clinical features. METHODS The histology findings were compared with patient demographics, clinical and renal parameters, mortality, CD4 count and date of commencing combined anti-retroviral therapy (cART). Follow-up was between 1 and 3.5 years. RESULTS We found a spectrum of renal histologies in HIV-positive patients of which HIV-associated nephropathy (HIVAN) was the most common histology. cART reduced the mortality in those with any feature of HIVAN by 57% [adjusted hazard ratio (AHR) 0.43, 95% confidence interval (CI) 0.22-0.85]. Of those patients with HIVAN who died, 79% died of renal failure as registered on their death certificate. Proteinuria and microcysts were shown to be poor prognostic indicators (AHR 1.36: 1.09-1.70 and 2.04: 1.24-3.37). In patients with HIVAN alone followed for up to 2 years on cART, estimated glomerular filtration rate remained stable and there was a trend towards decreased proteinuria. cART improved survival in patients with isolated immune complex disease. CONCLUSIONS As mortality is improved in patients with any feature of HIVAN or isolated immune complex disease, cART should be initiated once any of these histological features are established. We believe the spectrum of disease that constitutes HIVAN needs to be more specifically defined. The ultimate outcome may be determined by the histological subtype.

Complement C3 Plays an Essential Role in the Control of Opportunistic Fungal Infections

ABSTRACT The innate recognition of fungal pathogens is a crucial first step in the induction of protective antifungal immunity. Complement is thought to be one key component in this process, facilitating fungal recognition and inducing early inflammation. However, the roles of the individual complement components have not been examined extensively. Here we have used mice lacking C3 to examine its role in immunity to opportunistic fungal pathogens and show that this complement component is essential for resistance to infections with Candida albicans and Candida glabrata. We demonstrate that the absence of C3 impairs fungal clearance but does not affect inflammatory responses. We also show that the presence of C3 contributes to mortality in mice challenged with very high doses of Saccharomyces cerevisiae, although these effects were found to be mouse strain dependent.

Outcome of patients with membranous lupus nephritis in Cape Town South Africa

BACKGROUND The kidney is one of the major target organs affected by systemic lupus erythematosus. Although proliferative forms of lupus nephritis (LN) occur more frequently than membranous LN (MLN), the latter appears to have a more favourable outcome. Only a few studies have reported the outcome of patients with MLN. METHODS A retrospective analysis of patients with biopsy-confirmed MLN from a single centre in South Africa treated from 1st January 2000 to 31st December 2009. RESULTS The mean age of the patients (n = 42) at onset of LN was 35.0 ± 12.8 years with 73.8% of the patients being of mixed ancestry (coloureds). Eleven patients (26.2%) reached the composite end point of death or end-stage renal disease or persistent doubling of serum creatinine. The overall median survival and median renal survival times were 82.3 ± 15.5 months (95% confidence interval 52.0-112.6) and 84.5 ± 15.0 months (55.1-113.8), respectively. Also, 5-year event-free survival and renal survival were 64 and 71%, respectively. On multivariate analysis, systolic blood pressure (BP) during follow-up (P = 0.029), diastolic BP during follow-up (P = 0.020) and attainment of complete remission at 6 months (P = 0.033) were factors associated with the composite end points. Although treatment with chloroquine was not significantly associated with the composite end points (P = 0.05), we found that patients who received chloroquine had better renal survival compared with those who did not (P = 0.007). CONCLUSIONS The outcome of patients with MLN in Cape Town is poorer than for similar patients reported from other centres across the world. Better BP control may significantly influence outcome of disease in these patients.

EVIDENCE OF POSTNATAL ECDYSIS IN SOUTHERN RIGHT WHALES, EUBALAENA AUSTRALIS

Abstract This paper presents evidence of the presence and subsequent loss of postnatal skin in an ecdysis-like process in southern right whales, Eubalaena australis. Individuals whose skin was noticeably uneven, spongy, broken, and often light gray in color formed ≥20% of right whale neonates seen on the South African coast on any day up to and including the 1st week of September. Thereafter ≥85% of calves were of the normal, smooth-skinned appearance. The 50% transition point between the 2 forms occurred on 31 August (95% CI 1.1 days), or about a week after birth. Histological analysis of skin from stranded neonates showed a definite cleavage plane in the midepidermis, the mechanical integrity of which was further compromised by low concentrations of desmosomes and intracellular filaments. We propose that focal edema develops between the cells and forms the cleavage plane, which eventually leads to separation of the outer epidermal cell layer (cf. spongiosis in humans). The movement from the intrauterine to the oceanic milieu, and the osmoregulatory consequences thereof, may be a catalytic factor for this process to occur. This ecdysis may have important consequences for the cyamid fauna of neonatal right whales.

The Evolution of Our Knowledge of HIV-Associated Kidney Disease in Africa

Human immunodeficiency virus (HIV) infection started in Africa circa 1930. South Africa has the highest prevalence rate in the world. Although reports of HIV-associated nephropathy (HIVAN) appeared in the early 1980s, the earliest report from sub-Saharan Africa (SSA) came in 1994. Geographical, socioeconomic, political, and ethical factors have worked in concert to shape the character of HIV disease as it is seen in SSA. Political leaders within SSA have, through their actions, significantly contributed to the incidence of HIV infection. Black females, who often face cultural suppression and disadvantage, have a higher prevalence of HIV than males. Too few studies and outcomes data have bedeviled the statistics in SSA in relation to HIVAN prevalence and its management. Much of what is written is approximation and anecdotal. The largest reliable biopsy series comes from the University of Cape Town, where a workable classification of HIVAN has been developed to enable standardization of terminology. Histologic and clinical prognostic indicators with outcomes have been evaluated using this classification. Patients with HIV who present with acute kidney injury appear to have mainly acute tubular necrosis due to sepsis, dehydration, and nephrotoxic drugs. Since the rollout of combination antiretroviral therapy, the extent of HIV infection and kidney disease continues to be modified and possibly retarded.

The treatment of carcinoma in situ and squamous cell carcinoma of the conjunctiva with fractionated strontium-90 radiation in a population with a high prevalence of HIV

Background This study explores the safety and efficacy of strontium 90 (Sr-90) brachytherapy as the sole adjuvant therapy for carcinoma in situ (CIS) and squamous cell carcinoma (SCC) of the conjunctiva in a high HIV prevalent area. Methods This is a retrospective case review of patients treated with 60 Gray Sr-90 brachytherapy in four divided doses after resection with a 2 mm margin and histological confirmation. Cryotherapy or alcohol debridement was not performed at the time of excision due to limited resources. Two plaque sizes, 8.5 mm and 18 mm, were used. Results Sixty-nine patients were treated and had a median follow-up of 27 months (range 6–127). Thirty-three (47.8%) were HIV-positive. CIS was present in 40.6% and SCC in 59.4%. The surgical margins were positive in 39 (56.5%). Twenty patients (29.0%) were treated with the 18 mm plaque and 49 (71.0%) with the 8.5 mm plaque. Eight (11.6%) patients developed a recurrence at a median of 5 months (range 2–40). Recurrences only occurred in patients treated with the 8.5 mm plaque (p=0.094). There was no significant effect of HIV status, positive margins or staging on the number of recurrences. Treatment side effects were a dry eye in five patients which was successfully managed with topical lubricants, and induced astigmatism of 1 dioptre of cylinder in one patient. Conclusions Sr-90 brachytherapy is safe and effective in preventing recurrences in ocular surface squamous neoplasia in a high HIV prevalent setting. The 18 mm plaque size is superior to the 8.5 mm plaque size.

Outcome of patients with primary immune-complex type mesangiocapillary glomerulonephritis (MCGN) in Cape Town South Africa.

Background and Aim Mesangiocapillary glomerulonephritis (MCGN) is a common cause of chronic kidney disease in developing countries. Data on the renal outcome of patients with idiopathic MCGN is limited. The aim of this study is to investigate the outcome of patients with idiopathic MCGN presenting to the Groote Schuur Hospital (GSH) Renal Unit in Cape Town. Materials and Methods A retrospective study of patients with idiopathic MCGN followed up at our clinic. Seventy-nine patients with no identifiable cause of MCGN were included for analysis. A composite renal outcome of persistent doubling of serum creatinine or end stage renal disease (ESRD) was used. Kaplan Meier survival and Cox regression analysis were used to assess survival and identify factors predicting the outcome. Results The mean age at biopsy was 33.9±13.6 years and 41.8% were black. Mean duration of follow up was 13.5±18.8 months. Twenty-three patients (34.2%) reached the composite endpoint. Overall, median renal survival was 38.7±11.7 months (95% CI 15.7–61.8) with 2-year and 5-year renal survival of 61% and 40.3% respectively. No significant difference was found for renal survival between males and females, treatment or non-treatment with immunosuppression, presence or absence of crescents or histological type of MCGN (p>0.05). On univariate Cox-regression analysis, factors found to be associated with the outcome were the estimated glomerular filtration rate at biopsy (OR 0.97 [95%CI: 0.95–0.99], p<0.0001), black race (OR 3.03 [95%CI: 1.27–7.21], p = 0.012) and presence of interstitial fibrosis in the biopsy (OR 2.64 [95%CI: 1.07–6.48], p = 0.034). Age, systolic blood pressure and attaining complete or partial remission approached significant values with the endpoint. Conclusions The outcome of idiopathic MCGN in Cape Town is poor and requires further prospective studies to improve our understanding of this common disease.