Identification of HPV genotypes causing cervical precancer using tissue-based genotyping.

Identification of high-risk human papillomavirus genotypes causing cervical precancer is crucial for informing HPV vaccine development and efficacy studies, and for determining which types to include in next-generation genotyping assays. Co-occurrence of hrHPV infections is common and complicates carcinogenicity assessment; accurate attribution requires tissue-based genotyping of precancers. We included all women with cervical intraepithelial neoplasia Grade 2 or worse (CIN2+) from the Biopsy Study, an observational study of 690 women enrolled between 2009 and 2012 at the University of Oklahoma. Tissue-based genotyping, including whole tissue sections (WTS) and laser-capture microdissection (LCM), was performed on all precancers with multiple hrHPV infections detected in cytology, totaling over 1,800 HPV genotyping assays. Genotype attribution was compared to hierarchical and proportional hrHPV-type attribution models. Of 276 women with CIN2+, 122 (44.2%) had multiple hrHPV genotypes in cytology. Of 114 women with genotyping data, 94 had one or more hrHPV detected in tissue. Seventy-one women (75.5%) had a single causal hrHPV genotype, while 23 women had multiple hrHPV genotypes causing CIN2+. Ten women had multiple causal infections in a single biopsy, contrary to the previous notion that each lesion is caused by a single type only. While HPV16 was the predominant causal hrHPV genotype using all approaches, the hierarchical model overattributed HPV16, whereas other causal hrHPV genotypes, particularly HPV18 and HPV35, were underattributed. Understanding true causal genotypes is important for the evaluation of vaccine efficacy, to estimate the extent of unmasking, and for type-specific risk assessment in screening and management.

Renal biopsy findings among Indigenous Australians: a nationwide review.

Australia's Indigenous people have high rates of chronic kidney disease and kidney failure. To define renal disease among these people, we reviewed 643 renal biopsies on Indigenous people across Australia, and compared them with 249 biopsies of non-Indigenous patients. The intent was to reach a consensus on pathological findings and terminology, quantify glomerular size, and establish and compare regional biopsy profiles. The relative population-adjusted biopsy frequencies were 16.9, 6.6, and 1, respectively, for Aboriginal people living remotely/very remotely, for Torres Strait Islander people, and for non-remote-living Aboriginal people. Indigenous people more often had heavy proteinuria and renal failure at biopsy. No single condition defined the Indigenous biopsies and, where biopsy rates were high, all common conditions were in absolute excess. Indigenous people were more often diabetic than non-Indigenous people, but diabetic changes were still present in fewer than half their biopsies. Their biopsies also had higher rates of segmental sclerosis, post-infectious glomerulonephritis, and mixed morphologies. Among the great excess of biopsies in remote/very remote Aborigines, females predominated, with younger age at biopsy and larger mean glomerular volumes. Glomerulomegaly characterized biopsies with mesangiopathic changes only, with IgA deposition, or with diabetic change, and with focal segmental glomerulosclerosis (FSGS). This review reveals great variations in biopsy rates and findings among Indigenous Australians, and findings refute the prevailing dogma that most indigenous renal disease is due to diabetes. Glomerulomegaly in remote/very remote Aboriginal people is probably due to nephron deficiency, in part related to low birth weight, and probably contributes to the increased susceptibility to kidney disease and the predisposition to FSGS.

Estimating individual glomerular volume in the human kidney: clinical perspectives.

BACKGROUND: Measurement of individual glomerular volumes (IGV) has allowed the identification of drivers of glomerular hypertrophy in subjects without overt renal pathology. This study aims to highlight the relevance of IGV measurements with possible clinical implications and determine how many profiles must be measured in order to achieve stable size distribution estimates. METHODS: We re-analysed 2250 IGV estimates obtained using the disector/Cavalieri method in 41 African and 34 Caucasian Americans. Pooled IGV analysis of mean and variance was conducted. Monte-Carlo (Jackknife) simulations determined the effect of the number of sampled glomeruli on mean IGV. Lin's concordance coefficient (R(C)), coefficient of variation (CV) and coefficient of error (CE) measured reliability. RESULTS: IGV mean and variance increased with overweight and hypertensive status. Superficial glomeruli were significantly smaller than juxtamedullary glomeruli in all subjects (P < 0.01), by race (P < 0.05) and in obese individuals (P < 0.01). Subjects with multiple chronic kidney disease (CKD) comorbidities showed significant increases in IGV mean and variability. Overall, mean IGV was particularly reliable with nine or more sampled glomeruli (R(C) > 0.95, <5% difference in CV and CE). These observations were not affected by a reduced sample size and did not disrupt the inverse linear correlation between mean IGV and estimated total glomerular number. CONCLUSIONS: Multiple comorbidities for CKD are associated with increased IGV mean and variance within subjects, including overweight, obesity and hypertension. Zonal selection and the number of sampled glomeruli do not represent drawbacks for future longitudinal biopsy-based studies of glomerular size and distribution.

Distribution of volumes of individual glomeruli in kidneys at autopsy: association with physical and clinical characteristics and with ethnic group.

We have demonstrated considerable variability in the volumes of different glomeruli in given individuals (individual glomerular volume: IGV) in a stereologic study of kidneys at forensic autopsy performed to investigate sudden or unexpected death in people without manifest kidney disease. We review some important associations of IGV by subject characteristics and by ethnic groups. IGVs were measured by the Cavalieri method in 30 glomeruli in each of 111 adult males who belonged to 4 ethnic groups, i.e. US Whites, African-Americans, Africans from Senegal, and Australian Aborigines. Correlations of pooled IGV values with certain subject characteristics were evaluated in the US Whites. Pooled IGV data were compared in subjects across the 4 ethnic groups. In US Whites, mean IGV and its variance were greater with higher age, lower nephron number, lower birth weight, and with gross obesity, hypertension and cardiovascular death. In comparisons by ethnic group, mean IGV and IGV ranges were higher in African-Americans and Australian Aborigines than in US Whites and African Senegalese subjects. We conclude that glomerular enlargement with volume heterogeneity marks more advanced age, relative nephron deficiency, lower birth weight, obesity, hypertension, and advanced cardiovascular disease. The findings in African-Americans and Australian Aborigines suggest that larger IGVs and volume heterogeneity might mark populations with accentuated susceptibility to hypertension and kidney disease, but the data need to be further examined in the context of the determining characteristics defined in the US Whites.

Primer site polymorphisms: potential implications for bone marrow engraftment monitoring.

Assessment of chimerism after allogeneic bone marrow transplant frequently relies on PCR amplification of donor- and recipient-specific polymorphic short tandem repeats (STR) with subsequent fluorescent detection of amplicons. In vitro amplification of STR loci is often achieved through commercially available, multiplex PCR kits. While originally developed for forensic purposes, these kits are increasingly being used in clinical laboratories for early detection of graft failure and relapse of disease. Despite the obvious benefits of sensitivity, accuracy, reproducibility, and ease-of-use of these commercial kits, laboratories must understand the technological shortcomings of such assays to avoid misinterpretation of patient results. The current case illustrates how primer site polymorphisms associated with specific STR alleles can potentially lead to erroneous interpretation of engraftment analysis using one of these multiplex STR kits.

Nephron number, glomerular volume, renal disease and hypertension.

PURPOSE OF REVIEW: To discuss studies evaluating associations of glomerular number (Nglom) and glomerular volume with hypertension and kidney disease. IMPORTANT FINDINGS: The association of low Nglom with hypertension and renal insufficiency was described in the 1930s. Many investigators have noted loss of glomeruli with age, with most disappearing entirely, and have proposed that hypertension follows. In a recent German study, hypertensive patients had fewer glomeruli and larger mean glomerular volumes than nonhypertensive people. Among the 10-fold range of Nglom in our multiracial autopsy series, the lowest were in Australian Aborigines, who have the highest rates of renal failure. Nglom fell with age. There was a five-fold range in mean glomerular volume and considerable heterogeneity in individual glomerular volumes within a patient. Larger mean glomerular volume and greater individual glomerular volume heterogeneity correlated with lower Nglom, larger body size, hypertension, and black race. Hypertension increased with age and was marked by glomerular enlargement, intimal thickening and higher rates of glomerulosclerosis. In whites and Aborigines, but not in US blacks, lower Nglom was associated with hypertension, while robust numbers were highly protective. SUMMARY: Higher mean glomerular volume and individual glomerular volume heterogeneity mark glomerular stress. Low Nglom is an important determinant of hypertension and renal disease. Many 'missing' nephrons have probably been lost during life, leaving little trace. Additional factors contribute to high rates of hypertension in blacks.

Glomerular volume and clinicopathologic features related to disease severity in renal biopsies of African Americans and whites in the southeastern United States.

CONTEXT: African Americans have a 4-fold greater risk than whites for developing end-stage renal disease. Glomerulomegaly, possibly related to obesity, has been identified in high-risk populations and is suggested to be a marker for end-stage renal disease risk. OBJECTIVE: To investigate differences in glomerular size and patient clinical characteristics at the time of renal biopsy for the major diseases contributing to end-stage renal disease. DESIGN: Mean glomerular tuft volumes were estimated by the Weibel-Gomez method (1964) in native renal biopsies of 203 African American and 100 white patients 18 years of age and older by point counting on a stereologic grid. Glomerulosclerosis was graded on individual glomeruli from 0 to 4, and a glomerular sclerosis index was calculated for each biopsy. Relationships between the mean volume of nonsclerotic glomeruli, age, sex, race, sclerosis index, cortical fibrosis, estimated glomerular filtration rate, body mass index, and disease diagnosis were analyzed. RESULTS: Racial differences in mean volume of nonsclerotic glomeruli and body mass index were not significant in any disease category, and African Americans had more severe disease as determined by sclerosis index, cortical fibrosis, and estimated glomerular filtration rate only in focal segmental glomerulosclerosis. For all patients, increased sclerosis index and cortical fibrosis and lower estimated glomerular filtration rate were best predicted by increased age (P < .001). CONCLUSIONS: For approximately the same severity of disease, African Americans were 10 years or more younger than whites with the difference being seen in all disease categories except membranous glomerulonephritis and diabetes. Glomerulomegaly relative to whites was not a distinguishing feature of African American renal biopsies.

How many glomerular profiles must be measured to obtain reliable estimates of mean glomerular areas in human renal biopsies?

The objective of this study was to investigate the number of glomerular profiles that are required for accurate estimates of mean profile area in a renal biopsy series. Slides from 384 renal biopsies from one center were reviewed. They contained a median of seven glomerular profiles or of four profiles without sclerosis. Profile areas were measured using stereologic point counting. The "true individual mean" for each biopsy was calculated and the "true population mean" for groups of biopsies derived. Individual and population "random sample means" then were calculated from a random sampling of profiles in each biopsy and were compared with true means for the same biopsies. The effect on the true population means of the entire group of biopsies was also assessed, as the minimum number of glomerular profiles that were required for inclusion was changed. In a single biopsy, random sampling of > or =10 profiles without exclusions and of eight profiles or more without sclerosis reliably estimated the true mean areas. In a group of 30 biopsies, random sampling of five or more glomeruli per biopsy reliably estimated the true population mean. In the aggregate series, inclusion of all 384 biopsies produced the most robust true population mean; the reliability of the estimates decreased as the numbers of eligible biopsies diminished with increasing requisite minimum numbers of profiles per biopsy. We conclude that, while > or =10 profiles might be needed for reliable area estimates in a single biopsy, far fewer profiles per biopsy can suffice when groups of biopsies are studied. In analyses of groups of biopsies, all available biopsies should be used without consideration of the number of glomerular profiles in each. Stipulation of a specific minimum number of glomeruli in each biopsy for inclusion reduces the power of analyses because fewer biopsies are available for evaluation.