scholarly journals Digital Spatial Profiling of Collapsing Glomerulopathy

2021 ◽  
Author(s):  
Kelly D. Smith ◽  
Kammi Henriksen ◽  
Roberto F. Nicosia ◽  
Charles E. Alpers ◽  
Shreeram Akilesh
Keyword(s):  
Kidney Biopsy ◽  
Viral Infections ◽  
Transcriptional Profiling ◽  
Heavy Proteinuria ◽  
Collapsing Glomerulopathy ◽  
Spatial Profiling ◽  
Good Concordance ◽  
Laser Capture ◽  
Kidney Lesions

ABSTRACTCollapsing glomerulopathy is a histologically distinct variant of focal and segmental glomerulosclerosis that presents with heavy proteinuria and portends a poor prognosis. Collapsing glomerulopathy can be triggered by viral infections such as HIV and SARS-CoV-2. Transcriptional profiling of collapsing glomerulopathy lesions is difficult since only a few glomeruli may exhibit this histology within a kidney biopsy and the mechanisms driving this heterogeneity are unknown. Therefore, we used recently developed digital spatial profiling (DSP) technology which permits quantification of mRNA at the level of individual glomeruli. Using DSP, we profiled 1,852 transcripts in glomeruli from HIV and SARS-CoV-2 infected patients with biopsy confirmed collapsing glomerulopathy. The increased resolution of DSP uncovered heterogeneity in glomerular transcriptional profiles that were missed in early laser capture microdissection studies of pooled glomeruli. Focused validation using immunohistochemistry and RNA in situ hybridization showed good concordance with DSP results. Therefore, DSP represents a powerful method to dissect transcriptional programs of pathologically discernible kidney lesions.

Detection of Nucleic Acids in Cells or Tissue Sections Using In situ Polymerase Chain Reaction (PCR)

1996 ◽  
Vol 54 ◽  
pp. 16-17
Author(s):  
J. R. Hully ◽  
K. R. Luehrsen ◽  
K. Aoyagi ◽  
C. Shoemaker ◽  
R. Abramson
Keyword(s):  
Nucleic Acids ◽  
Viral Infections ◽  
Anatomical Location ◽  
Rt Pcr ◽  
Reverse Transcription Pcr ◽  
Cellular Source ◽  
Gene Transcripts ◽  
Initial Description ◽  
In Cells

The development of PCR technology has greatly accelerated medical research at the genetic and molecular levels. Until recently, the inherent sensitivity of this technique has been limited to isolated preparations of nucleic acids which lack or at best have limited morphological information. With the obvious exception of cell lines, traditional PCR or reverse transcription-PCR (RT-PCR) cannot identify the cellular source of the amplified product. In contrast, in situ hybridization (ISH) by definition, defines the anatomical location of a gene and/or it’s product. However, this technique lacks the sensitivity of PCR and cannot routinely detect less than 10 to 20 copies per cell. Consequently, the localization of rare transcripts, latent viral infections, foreign or altered genes cannot be identified by this technique. In situ PCR or in situ RT-PCR is a combination of the two techniques, exploiting the sensitivity of PCR and the anatomical definition provided by ISH. Since it’s initial description considerable advances have been made in the application of in situ PCR, improvements in protocols, and the development of hardware dedicated to in situ PCR using conventional microscope slides. Our understanding of the importance of viral latency or viral burden in regards to HIV, HPV, and KSHV infections has benefited from this technique, enabling detection of single viral copies in cells or tissue otherwise thought to be normal. Clearly, this technique will be useful tool in pathobiology especially carcinogenesis, gene therapy and manipulations, the study of rare gene transcripts, and forensics.

scholarly journals Specific sequences of infectious challenge lead to secondary hemophagocytic lymphohistiocytosis-like disease in mice

2019 ◽  
Vol 116 (6) ◽  
pp. 2200-2209 ◽  
Author(s):  
Andrew Wang ◽  
Scott D. Pope ◽  
Jason S. Weinstein ◽  
Shuang Yu ◽  
Cuiling Zhang ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Hemophagocytic Lymphohistiocytosis ◽  
Viral Infections ◽  
Transcriptional Profiling ◽  
Tlr4 Agonist ◽  
Tlr Agonist ◽  
Therapeutic Administration ◽  
Specific Sequences ◽  
Secondary Hemophagocytic Lymphohistiocytosis

Secondary hemophagocytic lymphohistiocytosis (sHLH) is a highly mortal complication associated with sepsis. In adults, it is often seen in the setting of infections, especially viral infections, but the mechanisms that underlie pathogenesis are unknown. sHLH is characterized by a hyperinflammatory state and the presence hemophagocytosis. We found that sequential challenging of mice with a nonlethal dose of viral toll-like receptor (TLR) agonist followed by a nonlethal dose of TLR4 agonist, but not other permutations, produced a highly lethal state that recapitulates many aspects of human HLH. We found that this hyperinflammatory response could be recapitulated in vitro in bone marrow-derived macrophages. RNA sequencing analyses revealed dramatic up-regulation of the red-pulp macrophage lineage-defining transcription factor SpiC and its associated transcriptional program, which was also present in bone marrow macrophages sorted from patients with sHLH. Transcriptional profiling also revealed a unique metabolic transcriptional profile in these macrophages, and immunometabolic phenotyping revealed impaired mitochondrial function and oxidative metabolism and a reliance on glycolytic metabolism. Subsequently, we show that therapeutic administration of the glycolysis inhibitor 2-deoxyglucose was sufficient to rescue animals from HLH. Together, these data identify a potential mechanism for the pathogenesis of sHLH and a potentially useful therapeutic strategy for its treatment.

scholarly journals Indirect effects of severe acute respiratory syndrome coronavirus 2 on the kidney in coronavirus disease patients

2020 ◽  
Vol 13 (3) ◽  
pp. 347-353 ◽  
Author(s):  
Aymeric Couturier ◽  
Sophie Ferlicot ◽  
Kévin Chevalier ◽  
Matthieu Guillet ◽  
Marie Essig ◽  
...  
Keyword(s):  
Severe Acute Respiratory Syndrome ◽  
Kidney Tissue ◽  
Kidney Injury ◽  
Renal Tubular Cells ◽  
Collapsing Glomerulopathy ◽  
Tubulointerstitial Lesions ◽  
Polymerase Chain ◽  
Renal Tubular ◽  
Novel Coronavirus ◽  
Kidney Lesions

Abstract Among patients hospitalized for novel coronavirus disease (COVID-19), between 10 and 14% develop an acute kidney injury and around half display marked proteinuria and haematuria. Post-mortem analyses of COVID-19 kidney tissue suggest that renal tubular cells and podocytes are affected. Here we report two cases of collapsing glomerulopathy and tubulointerstitial lesions in living COVID-19 patients. Despite our use of sensitive reverse transcription polymerase chain reaction techniques in this study, we failed to detect the virus in blood, urine and kidney tissues. Our observations suggest that these kidney lesions are probably not due to direct infection of the kidney by severe acute respiratory syndrome coronavirus 2.

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