scholarly journals Multi-Photon Microscopy for the Evaluation of Interstitial Fibrosis in Extended Criteria Donor Kidneys: A Proof-of-Concept Study

2021 ◽  
Author(s):  
Wei Zheng So ◽  
Rachel Zui Chih Teo ◽  
Li Yin Ooi ◽  
Benjamin Yen Seow Goh ◽  
Jirong Lu ◽  
...  
Keyword(s):  
Interstitial Fibrosis ◽  
Second Harmonic ◽  
Region Of Interest ◽  
Clinical Information ◽  
Average Score ◽  
Label Free ◽  
Tubular Atrophy ◽  
Photon Excitation ◽  
Multiple Biopsies ◽  
Auto Fluorescence

Abstract Introduction:This study evaluated the initial use of label-free second harmonic generation (SHG) imaging with two-photon excitation (2PE) auto-fluorescence for the quantification of collagen/fibrosis on pre-implantation biopsies of extended criteria donors (ECD).Methods:A total of 20 core ECD kidney transplant biopsy specimen tissues from 7 ECD donors, taken at the time of pre-implantation were retrieved, cut into 5-micron sections, mounted on slides and deparaffinized. The core needle biopsies were imaged with 2X and 20X objective using the commercially available laser-based Genesis® 200 (Histoindex Pte Ltd and Clinnovate Pte Ltd, Singapore). The entire core was selected as the Region of Interest (ROI). Corresponding clinical information from transplant donors were retrieved. Histopathological review was performed, and all biopsies had Interstitial Fibrosis (IF)/Tubular Atrophy (TA) scores > 0. Collagen parameters measured included quantification by the Collagen Area Ratio in Tissue (CART) and qualitative measurements by Collagen Reticulation Index (CRI).Results: 20 explant core biopsies were extracted from 10 donor kidney samples, of which originated from 7 donors. Table 1 depicts the baseline ECD characteristics of the donors. For the kidneys with multiple biopsies done, we obtained an average score of the collagen parameters across the different samples. Biopsies classified with > 85% KDPI score had significantly higher CAR and CART than biopsies with ≤ 85% KDPI score.Conclusion:MPM is an evolving technology that enables the quantification of the amount (CART) and quality (CRI) of collagen deposition in unstained explant biopsies of ECD kidneys. This initial evaluation found significant differences in both parameters between ECD kidneys with more or less than 85% KDPI scores.

scholarly journals Multi-Photon Microscopy for the Evaluation of Interstitial Fibrosis in Extended Criteria Donor Kidneys: A Proof-of-Concept Study

2021 ◽  
Author(s):  
Wei Zheng So ◽  
Rachel Zui Chih Teo ◽  
Li Yin Ooi ◽  
Benjamin Yen Seow Goh ◽  
Jirong Lu ◽  
...  
Keyword(s):  
Interstitial Fibrosis ◽  
Second Harmonic ◽  
Region Of Interest ◽  
Clinical Information ◽  
Average Score ◽  
Label Free ◽  
Tubular Atrophy ◽  
Photon Excitation ◽  
Multiple Biopsies ◽  
Auto Fluorescence

Abstract Introduction:This study evaluated the initial use of label-free second harmonic generation (SHG) imaging with two-photon excitation (2PE) auto-fluorescence for the quantification of collagen/fibrosis on pre-implantation biopsies of extended criteria donors (ECD).Methods:A total of 22 core ECD kidney transplant biopsy specimen tissues from 7 ECD donors, taken at the time of pre-implantation were retrieved, cut into 5-micron sections, mounted on slides and deparaffinized. The core needle biopsies were imaged with 2X and 20X objective using the commercially available laser-based Genesis® 200 (Histoindex Pte Ltd and Clinnovate Pte Ltd, Singapore). The entire core was selected as the Region of Interest (ROI). Corresponding clinical information from transplant donors were retrieved. Histopathological review was performed, and all biopsies had Interstitial Fibrosis (IF)/Tubular Atrophy (TA) scores > 0. Collagen parameters measured included quantification by the Collagen Area Ratio in Tissue (CART) and qualitative measurements by Collagen Reticulation Index (CRI).Results: 20 explant core biopsies were extracted from 10 donor kidney samples, of which originated from 7 donors. Table 1 depicts the baseline ECD characteristics of the donors. For the kidneys with multiple biopsies done, we obtained an average score of the collagen parameters across the different samples. Biopsies classified with > 85% KDPI score had significantly higher CAR and CART than biopsies with ≤ 85% KDPI score.Conclusion:MPM is an evolving technology that enables the quantification of the amount (CART) and quality (CRI) of collagen deposition in unstained explant biopsies of ECD kidneys. This initial evaluation found significant differences in both parameters between ECD kidneys with more or less than 85% KDPI scores.

scholarly journals Diagnosing Hirschsprung disease by detecting intestinal ganglion cells using label-free hyperspectral microscopy

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Marcos A. Soares de Oliveira ◽  
Laura Galganski ◽  
Sarah Stokes ◽  
Che -Wei Chang ◽  
Christopher D. Pivetti ◽  
...  
Keyword(s):  
Ganglion Cells ◽  
Second Harmonic ◽  
Hirschsprung Disease ◽  
Principal Component ◽  
Distal Colon ◽  
Support Vector ◽  
Label Free ◽  
Accurate Identification ◽  
Linear Discriminant ◽  
Multiple Biopsies

AbstractHirschsprung disease (HD) is a congenital disorder in the distal colon that is characterized by the absence of nerve ganglion cells in the diseased tissue. The primary treatment for HD is surgical intervention with resection of the aganglionic bowel. The accurate identification of the aganglionic segment depends on the histologic evaluation of multiple biopsies to determine the absence of ganglion cells in the tissue, which can be a time-consuming procedure. We investigate the feasibility of using a combination of label-free optical modalities, second harmonic generation (SHG); two-photon excitation autofluorescence (2PAF); and Raman spectroscopy (RS), to accurately locate and identify ganglion cells in murine intestinal tissue without the use of exogenous labels or dyes. We show that the image contrast provided by SHG and 2PAF signals allows for the visualization of the overall tissue morphology and localization of regions that may contain ganglion cells, while RS provides detailed multiplexed molecular information that can be used to accurately identify specific ganglion cells. Support vector machine, principal component analysis and linear discriminant analysis classification models were applied to the hyperspectral Raman data and showed that ganglion cells can be identified with a classification accuracy higher than 95%. Our findings suggest that a near real-time intraoperative histology method can be developed using these three optical modalities together that can aid pathologists and surgeons in rapid, accurate identification of ganglion cells to guide surgical decisions with minimal human intervention.

Identifying patients with CKD risk at the time of nephrectomy: When to initiate nephrology consult

2021 ◽  
pp. 239936932110319
Author(s):  
Yihe Yang ◽  
Zachary Kozel ◽  
Purva Sharma ◽  
Oksana Yaskiv ◽  
Jose Torres ◽  
...  
Keyword(s):  
Risk Factors ◽  
Multivariate Analysis ◽  
Radical Nephrectomy ◽  
Mixed Model ◽  
Linear Mixed Model ◽  
Interstitial Fibrosis ◽  
Kidney Neoplasm ◽  
Tubular Atrophy ◽  
Independent Risk Factors

Introduction: The prevalence of chronic kidney disease (CKD) is high among kidney neoplasm patients because of the overlapping risk factors. Our purpose is to identify kidney cancer survivors with higher CKD risk. Methods: We studied a retrospective cohort of 361 kidney tumor patients with partial or radical nephrectomy. Linear mixed model was performed. Results: Of patients with follow-up >3 months, 84% were identified retrospectively to fulfill criteria for CKD diagnosis, although CKD was documented in only 15%. Urinalysis was performed in 205 (57%) patients at the time of nephrectomy. Multivariate analysis showed interstitial fibrosis and tubular atrophy (IFTA) >25% ( p = 0.005), severe arteriolar sclerosis ( p = 0.013), female gender ( p = 0.024), older age ( p = 0.012), BMI ⩾ 25 kg/m2 ( p < 0.001), documented CKD ( p < 0.001), baseline eGFR ⩽ 60 ml/min/1.73 m2 ( p < 0.001), and radical nephrectomy ( p < 0.001) were independent risk factors of lower eGFR at baseline and during follow-up. Average eGFR decreased within 3 months post nephrectomy. However, patients with different risk levels showed different eGFR time trend pattern at longer follow-ups. Multivariate analysis of time × risk factor interaction showed BMI, radical nephrectomy and baseline eGFR had time-dependent impact. BMI ⩾ 25 kg/m2 and radical nephrectomy were associated with steeper eGFR decrease slope. In baseline eGFR > 90 ml/min/1.73 m2 group, eGFR rebounded to pre-nephrectomy levels during extended follow-up. In partial nephrectomy patients with baseline eGFR ⩾ 90 ml/min/1.73 m2 ( n = 61), proteinuria ( p < 0.001) and BMI ( p < 0.001) were independent risk factors of decreased eGFR during follow up. Conclusions: As have been suggested by others and confirmed by our study, proteinuria and CKD are greatly under-recognized. Although self-evident as a minimum workup for nephrectomy patients to include SCr, eGFR, urinalysis, and proteinuria, the need for uniform applications of this practice should be reinforced. Non-neoplastic histology evaluation is valuable and should include an estimate of global sclerosis% (GS) and IFTA%. Patients with any proteinuria and/or eGFR ⩽ 60 at the time of nephrectomy or in follow-up with urologists, and/or >25% GS or IFTA, should be referred for early nephrology consultation.

Hypertonic Dextrose Stimulates Chondrogenic Cells to Deposit Collagen and Proliferate

Cartilage ◽  
2021 ◽  
pp. 194760352110145
Author(s):  
Elisha Johnston ◽  
Yi Kou ◽  
Jason Junge ◽  
Lin Chen ◽  
Andrew Kochan ◽  
...  
Keyword(s):  
Metabolic Activity ◽  
Polynomial Regression ◽  
Second Harmonic ◽  
In Vitro Study ◽  
Cartilage Regeneration ◽  
Collagen Deposition ◽  
Cellular Processes ◽  
Photon Excitation ◽  
Hypertonic Dextrose

Objective Hypertonic dextrose (HD) injections (prolotherapy) for osteoarthritis are reported to reduce pain. Cartilage regeneration is hypothesized as a mechanism. This in vitro study identifies an HD concentration that stimulates chondrogenic cells to increase metabolic activity and assesses whether this concentration affects collagen deposition and proliferation. Design ATDC5 chondrogenic cells were cultured in normoglycemic DMEM/F12 medium, treated with concentrations of HD (4-400 mM), and assessed with PrestoBlue. Advanced light microscopy was used to conduct live imaging of collagen deposition through second harmonic generation microscopy (SHG) and proliferation via 2-photon excitation microscopy. Proliferation was additionally assessed with hemocytometer counts. Results A linear regression model found that, relative to the 4 mM baseline control, cells treated with 200 mM had a higher mean absorbance ( P = 0.023) and cells treated with 250 mM were trending toward a higher mean absorbance ( P = 0.076). Polynomial regression interpolated 240 mM as producing the highest average absorbance. Hemocytometer counts validated 250 mM as stimulating proliferation compared with the 4 mM control ( P < 0.01). A concentration of 250 mM HD led to an increase in collagen deposition compared with that observed in control ( P < 0.05). This HD concentration also led to increases in proliferation of ATDC5 cells relative to that of control ( P < 0.001). Conclusions A 250 mM HD solution appears to be associated with increased metabolic activity of chondrocytes, increased collagen deposition, and increased chondrocyte proliferation. These results support clinical prolotherapy research suggesting that intra-articular HD joint injections reduce knee pain. Further study of HD and cellular processes is warranted.

scholarly journals Advanced Label-Free Laser Scanning Microscopy and Its Biological Imaging Application

2021 ◽  
Vol 11 (3) ◽  
pp. 1002
Author(s):  
Xue Wang ◽  
Xinchao Lu ◽  
Chengjun Huang
Keyword(s):  
Laser Scanning ◽  
Second Harmonic ◽  
Biological Tissues ◽  
Biological Imaging ◽  
Laser Scanning Microscopy ◽  
Scanning Microscopy ◽  
Localized Surface Plasmon ◽  
Label Free ◽  
Coherent Raman ◽  
Surface Plasmon Microscopy

By eliminating the photodamage and photobleaching induced by high intensity laser and fluorescent molecular, the label-free laser scanning microscopy shows powerful capability for imaging and dynamic tracing to biological tissues and cells. In this review, three types of label-free laser scanning microscopies: laser scanning coherent Raman scattering microscopy, second harmonic generation microscopy and scanning localized surface plasmon microscopy are discussed with their fundamentals, features and recent progress. The applications of label-free biological imaging of these laser scanning microscopies are also introduced. Finally, the performance of the microscopies is compared and the limitation and perspectives are summarized.

Label-free second-harmonic phase imaging of biological specimen by digital holographic microscopy

2010 ◽  
Vol 35 (24) ◽  
pp. 4102 ◽  
Author(s):  
Etienne Shaffer ◽  
Corinne Moratal ◽  
Pierre Magistretti ◽  
Pierre Marquet ◽  
Christian Depeursinge
Keyword(s):  
Second Harmonic ◽  
Digital Holographic Microscopy ◽  
Phase Imaging ◽  
Label Free ◽  
Biological Specimen ◽  
Harmonic Phase ◽  
Holographic Microscopy ◽  
Harmonic Phase Imaging

scholarly journals Cellular Mechanisms of Tissue Fibrosis. 3. Novel mechanisms of kidney fibrosis

2013 ◽  
Vol 304 (7) ◽  
pp. C591-C603 ◽  
Author(s):  
Gabriela Campanholle ◽  
Giovanni Ligresti ◽  
Sina A. Gharib ◽  
Jeremy S. Duffield
Keyword(s):  
Chronic Kidney Disease ◽  
Kidney Disease ◽  
Interstitial Fibrosis ◽  
Kidney Injury ◽  
Innate Immune ◽  
Cellular Mechanisms ◽  
Tubular Atrophy ◽  
Kidney Fibrosis ◽  
Capillary Rarefaction ◽  
Peritubular Capillaries

Chronic kidney disease, defined as loss of kidney function for more than three months, is characterized pathologically by glomerulosclerosis, interstitial fibrosis, tubular atrophy, peritubular capillary rarefaction, and inflammation. Recent studies have identified a previously poorly appreciated, yet extensive population of mesenchymal cells, called either pericytes when attached to peritubular capillaries or resident fibroblasts when embedded in matrix, as the progenitors of scar-forming cells known as myofibroblasts. In response to sustained kidney injury, pericytes detach from the vasculature and differentiate into myofibroblasts, a process not only causing fibrosis, but also directly contributing to capillary rarefaction and inflammation. The interrelationship of these three detrimental processes makes myofibroblasts and their pericyte progenitors an attractive target in chronic kidney disease. In this review, we describe current understanding of the mechanisms of pericyte-to-myofibroblast differentiation during chronic kidney disease, draw parallels with disease processes in the glomerulus, and highlight promising new therapeutic strategies that target pericytes or myofibroblasts. In addition, we describe the critical paracrine roles of epithelial, endothelial, and innate immune cells in the fibrogenic process.

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