Markers for the Study of Diabetic Neuropathy

Widely Used and Frequently Published
Our Pain and Inflammation Research Antibodies have proven valuable for the study of neuropathic, nociceptive and inflammatory pain.

This recent publication is focused on Diabetic Neuropathy. José Eduardo Roa-Coria, Jorge Baruch Pineda-Farias, Paulino Barragán-Iglesias, Geovanna Nallely Quiñonez-Bastidas, Ángel Zúñiga-Romero, Juan Carlos Huerta-Cruz, Juan Gerardo Reyes-García, Francisco Javier Flores-Murrieta, Vinicio Granados-SotoView ORCID ID profile and Héctor Isaac Rocha-González (2019). Possible involvement of peripheral TRP channels in the hydrogen sulfide-induced hyperalgesia in diabetic rats. BMC Neuroscience 201920:1 https://doi.org/10.1186/s12868-018-0483-3.

The results show Hydrogen Sulfide (H₂S) catalyzes hyperalgesia in rats via TRP Channels. Our Substance P is used in the study.

Figure: Immunolocalization of cystathionin-β-synthase enzyme (CBS, red) with a–e neuronal nuclear antigen (NeuN)-, f–j substance P (SP)-, k–o isolectin B4 (IB4)- and p–t glial fibrillary acidic protein (GFAP)-positive dorsal root ganglion neurons of diabetic rats (green). a, f, k and p show a representative 16 µm-slice from dorsal root ganglia. b, g, l and q show a representative CBS staining with Cy3 from dorsal root neurons, whereas c, h, m and r show NeuN, SP, IB4 and GFAP representative staining with Cy2 in dorsal root neurons, respectively. d, i, n and s show the merged image from Cy3 and Cy2 signals, the overlapping between CBS and neuronal markers is shown in a yellow-orange color. e, j, o and t show a magnification of d, i, n and s, respectively.

The authors conclude that H2S leads to hyperalgesia in diabetic rats through activation of TRPV1, TRPA1 and TRPC channels and, subsequent intraepidermal fibers loss. CBS enzyme inhibitors or TRP-channel blockers could be useful for the treatment of painful diabetic neuropathy.

TRP Antibodies

Some of the Best Available
Our TRP Channel Antibodies are widely used and frequently published.

Here's some data from researchers using on of our TRPV1 Antibodies.

Immunohistochemical expression of serum response factor in rat dorsal root ganglia. Dorsal root ganglia dissected from male Sprague-Dawley rat and imaged by confocal microscopy for (A) TRPV1 (blue), (B) AKAP (AKAP150, green), and (C) SRF (red), with colocalization represented in panel (D). White arrows indicate cells expressing all 3 proteins. Yellow line represents 50 μM in length; images are representative of 4 individual sections taken from 6 L4 to L6 DRG. (E) Venn diagram of coexpression between SRF (red circle), AKAP (AKAP150, green circle), and TRPV1 (blue circle). (F) Percentages of SRF coexpression with AKAP (AKAP150) or TRPV1 calculated from 8 randomly selected coverslips. DRG, dorsal root ganglia; SRF, serum response factor. Source Serum response factor mediates nociceptor inflammatory pain plasticity PAIN Reports3(3):e658, May/June 2018.

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TRPV1 and Diabetic Neuropathy

Thermal hyperalgesia is a common sympton of Diabetic Periperal Neuropathy (DPN). It is one of most difficult types of pain to treat. The development of tolerance, inadequate relief and potential toxicity of classical antinociceptives warrant the investigation of the newer agents to relieve this pain.

The elevated expression of Transient receptor potential vanilloid 1 (TRPV1) suspected as a transmitter of this pain. Dr. Louis Premkumar and his team at SIU have recently published results that further demonstrate the role of TRPV1: Mahendra Bishnoi, Christine A Bosgraaf, Mruvil Abooj, Linlin Zhong, Louis S Premkumar. Streptozotocin-Induced Early Thermal Hyperalgesia is independent of Glycemic State of Rats: Role of Transient Receptor Potential Vanilloid 1(TRPV1) and inflammatory mediators. Molecular Pain 2011, 7:52 doi:10.1186/1744-8069-7-52. Published: 27 July 2011.

Figure 4. Altered TRPV1 staining in spinal cord dorsal horn of STZ-treated rats. A. Representative images of TRPV1 staining from a vehicle-treated, STZ-HG and STZNG rats. An enlarged segment has also been shown. B. Average gray values/10,000 μm² area of TRPV1 staining in dorsal horn was significantly increased (p<0.05) in both STZ-HG and STZ-NG rats as compared to vehicle-treated rats. Asterisk (*) represents p < 0.05. Scale bar is 200 μm and 50 μm for upper and lower panels, respectively.

Conclusions: From these results, it is concluded that TRPV1 is an integral component of initiating and maintaining inflammatory thermal hyperalgesia, which can be alleviated by intrathecal administration of RTX. Further, the results suggest that enhanced expression and inflammation-induced sensitization of TRPV1 at the spinal cord may play a role in central sensitization in STZ-induced neuropathy.

Therapies that downregulate or silence TRPV1 expression could be the key to better treatments for the Thermal Algesia cause by diabetes. I will keep you posted.

TRPV1 and Retinal Ganglion Cell Apoptosis

The publications referencing our TRPV (Vanilloid); TRPM; TRPA and TRPC Antibodies keep on coming.

In this February 2009 publication, Dr. David Calkins and his team at Vanderbilt demontrate that Retinal Ganglian Cells express the TRPV1 channel and that TRPV1 activation contributes to their death with exposure to hydrostatic pressure . We also demonstrated that activation of TRPV1 alone was sufficient to induce apoptosis of RGCs.

Rebecca M. Sappington, Tatiana Sidorova, Daniel J. Long, and David J. Calkins. TRPV1: Contribution to Retinal Ganglion Cell Apoptosis and Increased Intracellular Ca2+ with Exposure to Hydrostatic Pressure. Invest. Ophthalmol. Vis. Sci., Feb 2009; 50: 717 - 728.
...we used rabbit anti-mouse TRPV1 IgG (1:500; catalog number RA14113; Neuromics, Edina, MN) against the absolute C terminus of mouse TRPV1 (EDAEVFKDSMAPGEK)...

TRPV1 localization in RGCs of rat retina. (A) Immunocytochemical labeling for TRPV1 shows strong localization in the outer retina and in RGCs (large cell bodies); there is little or no label in smaller displaced amacrine cells of the GCL. Clear examples of amoeboid-shaped cell bodies of microglia cells are indicated (ovals). Right: Control section preabsorbed using the TRPV1 blocking peptide (+BP). (B) Confocal image stack through GCL and NFL showing labeling for TRPV1 in wholemount preparation counterlabeled with antibodies against heavy-chain neurofilaments that recognize broad-field RGCs (SMI32). Image shows punctate localization to dendrites (arrows) as well as intense label to cell bodies (brackets); smaller cell bodies with TRPV1 label are in the background. (C) Confocal image stack through GCL and NFL of peripheral retina shows TRPV1 in RGC cell bodies (bracket) and in small bundles of RGC axons (arrows). (D) Confocal stack from central retina shows TRPV1 in RGC cell bodies (bracket) and in axon bundles in the NFL as they course toward the optic nerve head. Amoeboid-shaped cell bodies of microglia are apparent (ovals). (E) Confocal image in single plane at GCL/NFL border shows Iba-1–labeled microglia processes colocalizing with TRPV1, as we previously demonstrated.51 TRPV1-label RGC cell bodies (brackets) and axons (arrows) are indicated for reference. (F) Immunocytochemical labeling demonstrates strong perinuclear and dendritic localization of TRPV1 in cultured RGCs counterstained with the nuclear label DAPI. Localization to dendritic processes and neurites (dashed circles) includes node-like clusters; right: these regions are shown at higher magnification. (G, top) Western blot against TRPV1 in brain and whole retina from adult rat shows band at expected molecular weight (arrowheads; 100–113 kDa). Retina demonstrates an additional band with a slightly lower molecular weight that probably corresponds to a different glycosylation state for this antibody.79 Bottom: Control Western blot with preabsorption of TRPV1 antibody using the blocking peptide prevents detection of both bands. ONL, outer nuclear layer; OPL, outer plexiform layer; INL, inner nuclear layer; IPL, inner plexiform layer; GCL, ganglion cell layer; NFL, nerve fiber layer.

TRPV1 Staining of Mouse DRG

This excellent staining comes to us courtesy of Katharina Zimmermann (Childrens Hospital Boston, Clapham Lab). This is some of the best staining we've seen using our VR1 C-Terminus (TRPV1) - mouse specific antibody.

Images: TRPV1 staining of C57BL/6 mouse dorrsal root ganglia. Tissues were stained using Alexa Fluor© 488 (Green) and counterstained with DAPI (blue).

cryosections, 10 microns thickness