Optical clearing reveals TNBS-induced morphological changes of VGLUT2-positive nerve fibers in mouse colorectum

2021 ◽  
Author(s):  
Tiantian Guo ◽  
Shivam P Patel ◽  
Dhruv Shah ◽  
Ling Chi ◽  
Sharareh Emadi ◽  
...  
Keyword(s):  
Myenteric Plexus ◽  
Morphological Changes ◽  
Glutamate Transporter ◽  
Nerve Fibers ◽  
Trinitrobenzenesulfonic Acid ◽  
Fiber Density ◽  
Irritable Bowel ◽  
Almost All ◽  
Nerve Fiber Density

Colorectal hypersensitivity and sensitization of both mechanosensitive and mechanically insensitive afferents develop after intracolonic instillation of 2,4,6-trinitrobenzenesulfonic acid (TNBS) in the mouse, a model of post-infectious irritable bowel syndrome. In mice in which ~80% of extrinsic colorectal afferents were labeled genetically using the promotor for vesicular glutamate transporter type 2 (VGLUT2), we systematically quantified the morphology of VGLUT2-positive axons in mouse colorectum 7-28 days following intracolonic TNBS treatment. After removal, the colorectum was distended (20 mmHg), fixed with paraformaldehyde, and optically cleared to image VGLUT2-positive axons throughout the colorectal wall thickness. We conducted vector path tracing of individual axons to allow systematic quantification of nerve fiber density and shape. Abundant VGLUT2-positive nerve fibers were present in most layers of the colorectum, except the serosal and longitudinal muscular layers. A small percentage of VGLUT2-positive myenteric plexus neurons was also detected. Intracolonic TNBS treatment significantly reduced the number of VGLUT2-positive nerve fibers in submucosal, myenteric plexus, and mucosal layers at day 7 post TNBS, which mostly recovered by day 28. We also found that almost all fibers in the submucosa were meandering and curvy, with ~10% showing pronounced curviness (quantified by the linearity index). TNBS treatment resulted in a significant reduction of the proportions of pronounced curvy fibers in the rectal region at 28 days post TNBS. Altogether, the present morphological study reveals profound changes in the distribution of VGLUT2-positive fibers in mouse colorectum undergoing TNBS-induced colitis, and draw attention to curvy fibers in the submucosa with potential roles in visceral nociception.

scholarly journals Impact of Dose, Sex, and Strain on Oxaliplatin-Induced Peripheral Neuropathy in Mice

2021 ◽  
Vol 2 ◽  
Author(s):  
Urszula O. Warncke ◽  
Wisam Toma ◽  
Julie A. Meade ◽  
Abigail J. Park ◽  
Danielle C. Thompson ◽  
...  
Keyword(s):  
Peripheral Neuropathy ◽  
Nerve Conduction ◽  
Side Effect ◽  
Nerve Fibers ◽  
Therapeutic Interventions ◽  
Fiber Density ◽  
Intraepidermal Nerve Fiber Density ◽  
Nerve Fiber Density

Chemotherapy-induced peripheral neuropathy (CIPN) is a common, dose limiting, and long-lasting side effect of chemotherapy treatment. Unfortunately, no treatment has proven efficacious for this side effect. Rodent models play a crucial role in the discovery of new mechanisms underlying the initiation, progression, and recovery of CIPN and the potential discovery of new therapeutics. However, there is limited consistency in the dose, the sex, age, and genetic background of the animal used in these studies and the outcome measures used in evaluation of CIPN rely primarily on noxious and reflexive measures. The main objective of this study was to provide a comprehensive and systematic characterization of oxaliplatin-induced peripheral neuropathy in mice by using a battery of behavioral, sensory, electrophysiological, and morphometric measures in both sexes of the two widely used strains of mice, C57BL/6J and BALB/cJ. Mice received intraperitoneal injections of 3 or 30 mg/kg cumulative doses of oxaliplatin over the course of 2 weeks. Both doses induced long-term and time-dependent mechanical and cold hypersensitivity. Our results show that 30 mg/kg oxaliplatin reduced the locomotor activity in C57BL/6J mice, and C57BL/6J females showed anxiety-like behavior one-week post completion of treatment. In the same dose group, BALB/cJ males and females sustained a larger decrease in sucrose preference than either male or female C57BL/6J mice. Both strains failed to show significant changes in burrowing and nesting behaviors. Two clinically relevant assessments of changes to the peripheral nerve fibers, nerve conduction and intraepidermal nerve fiber density (IENFD) were evaluated. Only BALB/cJ females showed significant reduction in the nerve conduction amplitude 1 week after 30 mg/kg oxaliplatin regimen. Moreover, this dose of the chemo agent reduced the IENF density in both sexes and strains. Our findings suggest that mouse strain, sex, and assay type should be carefully considered when assessing the effects of oxaliplatin and potential therapeutic interventions.

scholarly journals Nerve fibers in the Tumor Microenvironment are co-localized with Tertiary Lymphoid Structures

2020 ◽  
Author(s):  
Lara R. Heij ◽  
Xiuxiang Tan ◽  
Jakob N. Kather ◽  
Jan M. Niehues ◽  
Shivan Sivakumar ◽  
...  
Keyword(s):  
Pancreatic Ductal Adenocarcinoma ◽  
Immune Cells ◽  
Immune Cell ◽  
Nerve Fibers ◽  
Ductal Adenocarcinoma ◽  
Fiber Density ◽  
Tertiary Lymphoid Structures ◽  
Lymphoid Structures ◽  
Nerve Fiber Density ◽  
Small Nerve

ABSTRACTBackgroundB cells and tertiary lymphoid structures (TLS) are reported to be important in the improvement of survival of cancer patients. These secondary lymphoid organs have been associated with the generation of an anti-tumor response. Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal cancer types and the stromal architecture shapes the intratumoral heterogeneity. The stroma of PDAC is a complex system in which crosstalk takes place between cancer-associated fibroblasts, immune cells, endothelial cells and the cancer cells. Besides immune cells and fibroblasts, there is some limited data about the influence of nerve fibers on cancer progression.Patients and methodsNerve Fiber Density (NFD) was analysed in our cohort of 188 patients with Pancreatic Ductal Adenocarcinoma who underwent pancreatic surgery. We used immunohistochemistry and multiplex imaging to phenotype the immune cell infiltrate. The cell detection classifier measured distance from immune cell to cancer gland and with a heat map we could count TLS. By using Machine learning we were able to define the spatial distribution and counting Tertiary Lymphoid Structures.ResultsHigh NFD is significantly associated with prolonged overall survival (HR 1.676 (95%CI 1.126,2.495) for low vs. high NFD, p-value 0.0109). The immune cells surrounding the nerve fibers were phenotyped in B cells, T cells and dendritic follicular cells, matching a TLS. Here we show that small nerve fibers are located at the TLS in Pancreatic Cancer and a high Nerve Fiber Density combined with more than 5 TLS is associated with a better survival (HR 0.388 (95%CI 0.218, 0.689).ConclusionThe co-localization of small nerve fibers with TLS is a new finding which has not been described before. However the precise roles of these TLS and nerve fibers remains unknown. These findings unravel future pathways and has the potential to reach new directions into already existing targeted therapy.

scholarly journals Intraepidermal Nerve fiber Density in healthy Subjects with Diabetic Family History. ¿Is Neuropathy an early Biomarker? A Pilot study

2021 ◽  
Vol 8 (3) ◽  
pp. 01-08
Author(s):  
Ildefonso Leyva
Keyword(s):  
Pilot Study ◽  
Family History ◽  
Nerve Fiber ◽  
Healthy Subjects ◽  
Nerve Fibers ◽  
Diabetic Patients ◽  
Fiber Density ◽  
Intraepidermal Nerve Fiber Density ◽  
Nerve Fiber Density ◽  
Intraepidermal Nerve Fiber

Objective: Evaluate the intraepidermal nerve fiber density in healthy subjects with diabetic family history compared with diabetic patients and controls. Introduction: Neuropathy is the most prevalent chronic complication of diabetes, presenting various symptoms that interfere with daily living activities, psychosocially disability, and reducing life quality. The skin biopsy is recognized as a minimally invasive procedure that allows morphometric quantification of intraepidermal nerve fibers and has made possible the study of peripheral neuropathies involving thin fibers that traditional methods cannot diagnose. Methods: Analytical cross-sectional observational pilot study with seven patients per group including healthy, diabetic, and healthy with diabetic family history subjects. For the statistical analysis, we used the R package, R software version 3.3.2, with a confidence level of 95%. The research was performed with ANOVA and Kruskal-Wallis test to test the primary objective. Results: The density of intraepidermal nerve fibers is similar between the group with diabetic family history 6.8 ± 2.1 (3.5 - 10.1) and diabetic patients 6.3 ± 2.9 (3.5 - 7.05) while the control group reported a density in parameters of normality of 10± 1.2 (8.2 - 10.1) with a p= 0.01 between the three groups. The decrease of intraepidermal nerve fibers showed a tendency to decrease with increasing age and BMI with a ratio coefficient for age of r= -0.342, 95% CI (-0.67 - 0.106), p= 0.129; and for BMI of r= -0.36, 95% CI (-0.685 - 0.0847), p= 0.109. Conclusion: Intraepidermal nerve fiber density is decreased in subjects with a family history of diabetes mellitus type 2 and even more so in diabetics, with no statistical difference.

566-P: Mild Increase of Proinflammatory Macrophages Correlates with Reduction of Nerve Fiber Density in Sural Nerve of Human Type 2 Diabetic Subjects

Diabetes ◽  
2019 ◽  
Vol 68 (Supplement 1) ◽  
pp. 566-P
Author(s):  
HIROKI MIZUKAMI ◽  
SHO OSONOI ◽  
KAZUHISA TAKAHASHI ◽  
KAZUHIRO KUDO ◽  
CHIEKO ITABASHI ◽  
...  
Keyword(s):  
Nerve Fiber ◽  
Sural Nerve ◽  
Fiber Density ◽  
Human Type ◽  
Type 2 Diabetic ◽  
Nerve Fiber Density

scholarly journals Long-term sensitization of mechanosensitive and -insensitive afferents in mice with persistent colorectal hypersensitivity

2012 ◽  
Vol 302 (7) ◽  
pp. G676-G683 ◽  
Author(s):  
Bin Feng ◽  
Jun-ho La ◽  
Erica S. Schwartz ◽  
Takahiro Tanaka ◽  
Timothy P. McMurray ◽  
...  
Keyword(s):  
Afferent Input ◽  
Significant Loss ◽  
Pelvic Nerve ◽  
Fiber Density ◽  
Electrophysiological Recordings ◽  
Visceromotor Response ◽  
Irritable Bowel ◽  
Nerve Fiber Density ◽  
Circumferential Stretch

Afferent input contributes significantly to the pain and colorectal hypersensitivity that characterize irritable bowel syndrome. In the present study, we investigated the contributions of mechanically sensitive and mechanically insensitive afferents (MIAs; or silent afferents) to colorectal hypersensitivity. The visceromotor response to colorectal distension (CRD; 15–60 mmHg) was recorded in mice before and for weeks after intracolonic treatment with zymosan or saline. After CRD tests, the distal colorectum with the pelvic nerve attached was removed for single-fiber electrophysiological recordings. Colorectal afferent endings were located by electrical stimulation and characterized as mechanosensitive or not by blunt probing, mucosal stroking, and circumferential stretch. Intracolonic zymosan produced persistent colorectal hypersensitivity (>24 days) associated with brief colorectal inflammation. Pelvic nerve muscular-mucosal but not muscular mechanosensitive afferents recorded from mice with colorectal hypersensitivity exhibited persistent sensitization. In addition, the proportion of MIAs (relative to control) was significantly reduced from 27% to 13%, whereas the proportion of serosal afferents was significantly increased from 34% to 53%, suggesting that MIAs acquired mechanosensitivity. PGP9.5 immunostaining revealed no significant loss of colorectal nerve fiber density, suggesting that the reduction in MIAs is not due to peripheral fiber loss after intracolonic zymosan. These results indicate that colorectal MIAs and sensitized muscular-mucosal afferents that respond to stretch contribute significantly to the afferent input that sustains hypersensitivity to CRD, suggesting that targeted management of colorectal afferent input could significantly reduce patients' complaints of pain and hypersensitivity.

scholarly journals Reduced Corneal Nerve Fiber Density in Type 2 Diabetes by Wide-Area Mosaic Analysis

2017 ◽  
Vol 58 (14) ◽  
pp. 6318 ◽  
Author(s):  
Neil S. Lagali ◽  
Stephan Allgeier ◽  
Pedro Guimarães ◽  
Reza A. Badian ◽  
Alfredo Ruggeri ◽  
...  
Keyword(s):  
Type 2 Diabetes ◽  
Nerve Fiber ◽  
Wide Area ◽  
Fiber Density ◽  
Mosaic Analysis ◽  
Corneal Nerve ◽  
Nerve Fiber Density

scholarly journals PD1+ T-cells correlate with Nerve Fiber Density as a prognostic biomarker in patients with resected perihilar cholangiocarcinoma.

2022 ◽  
Author(s):  
Xiuxiang Tan ◽  
Mika Rosin ◽  
Simone Appinger ◽  
Jan Bednarsch ◽  
Dong Liu ◽  
...  
Keyword(s):  
T Cells ◽  
Nerve Fiber ◽  
Immune Cell ◽  
Prognostic Biomarker ◽  
Region Of Interest ◽  
Nerve Fibers ◽  
Perihilar Cholangiocarcinoma ◽  
Fiber Density ◽  
Nerve Fiber Density ◽  
Small Nerve

Background & Aims: Perihilar cholangiocarcinoma (pCCA) is a hepatobiliary malignancy. Nerve fiber invasion (NFI) shows cancer invading the nerve and is considered an aggressive feature. Nerve fiber density (NFD) consists of small nerve fibers without cancer invasion and is divided into high NFD (high numbers of small nerve fibers) or low NFD (low numbers of small nerve fibers). We aim to explore differences in immune cell populations and survival. Approach & Results: We applied multiplex immunofluorescence (mIF) on 47 pCCA surgically resected patients and investigated immune cell composition in the tumor microenvironment (TME) of nerve fiber phenotypes (NFI, high and low NFD). Group comparison was performed and overall survival (OS) was assessed. The NFI Region of Interest (ROI) was measured with highest CD68+ macrophage levels among 3 ROIs (NFI compared to tumor free p= 0.016 and to tumor p=0.034) and PD1 expression on CD8 and were more abundant in the tumor rather than NFI ROI (p= 0.004 and p= 0.0029 respectively). NFD compared to NFI, demonstrated co-expression of CD8+PD1+ as well as CD68+PD1+ to be significantly higher in high NFD patients (p= 0.027 and p= 0.044, respectively). The high NFD OS was 92 months median OS (95% CI:41-142), for low NFD 20 months ((95% CI: 4-36) and for NFI 19 months (95% CI 7-33). High NFD OS was significantly better compared to low NFD (p= 0.046) and NFI (p= 0.032). Conclusions: PD1+ T-cells correlate with high NFD as a prognostic biomarker, the biological pathway behind this needs to be investigated.

scholarly journals Assessment and comparison of the morphology and function of the corneal sub-basal nerve plexus in type-1 diabetes mellitus patients and in healthy subjects

2016 ◽  
Vol 1 (1) ◽  
pp. 51-63
Author(s):  
Irmante Derkac ◽  
Ingrida Januleviciene ◽  
Kirwan Asselineau ◽  
Dzilda Velickiene
Keyword(s):  
Nerve Fiber ◽  
Healthy Subjects ◽  
Nerve Fibers ◽  
Corneal Sensitivity ◽  
Fiber Density ◽  
Corneal Confocal Microscopy ◽  
Corneal Nerve ◽  
Type 1 Dm ◽  
Nerve Fiber Density

Aim/purpose: It is believed that small nerve bundles are damaged in the earliest stages of neuropathy caused by diabetes mellitus (DM). Our goal was to evaluate and compare anatomical characteristics of corneal nerve fibers and corneal sensitivity in type-1 DM patients and in healthy control subjects.Design: A prospective, masked, controlled cross-sectional clinical study.Method: Thirty patients with type-1 DM and ten non-diabetic healthy subjects underwent a corneal confocal microscopy to evaluate the corneal sub-basal nerve fibers (density, number of nerves and branches, total nerve length) and contact corneal esthesiometry.Results: Diabetic patients had significantly lower corneal nerve fiber density density (14.32 ± 5.87 vs. 19.71 ± 5.59 mm/mm2; p = 0.023 ) nerve branches number (4.57 ± 3,91 vs. 9.90 ± 5.8 n°/image; p = 0.006) , nerve fiber length (2.28 ± 0.94 vs. 3.13 ± 0.89 mm; p = 0.032) and corneal sensitivity (1.13 ± 0.29 vs. 0.98 ± 0.058 gr/mm2 p = 0.02), as compared with controls. A negative correlation was found between corneal nerve fiber length, corneal nerve number, corneal nerve fiber density and disease duration (p < 0.05).Conclusion: Corneal confocal microscopy and corneal sensitivity evaluation are noninvasive techniques helping to detect early changes in the sub-basal nerve plexus characteristic for diabetic neuropathy (DN) in patients with type-1 DM. Further studies are required to investigate the role of corneal neuropathy assessment using these novel techniques as a toll to detect early DN. 

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