Journal: Molecular Pain

Article Title: Neurotrophic factor changes in the rat thick skin following chronic constriction injury of the sciatic nerve

doi: 10.1186/1744-8069-8-1

Figure Lengend Snippet: Distribution of p75 immunoreactivity following nerve injury and relationship to nerve fibers and proNGF . A) In sham-operated animals, p75 was distributed (red) around and with PGP 9.5-IR nerve fibers (green). P75 staining was found more clearly around large cutaneous PGP 9.5-IR nerve fiber bundles and smaller fibers along the dermo-epidermal junction. Where nerve fibers crossed the dermo-epidermal junction into the epidermis, the yellow color representing p75 associating with nerve fibers was lost (arrow). B) At 1 week post-injury, virtually all PGP-9.5-IR nerve fibers disappeared from the upper dermis and epidermis; immunostaining in p75-IR Schwann was very intense. C) At 2 weeks post-injury, a low number of PGP-9.5-IR fibers were detected and were associated with p75 Schwann cells (yellow), however most of the PGP-9.5-IR was restricted to Langerhans cells in epidermis (arrow) D) At 4 weeks post-injury, p75 immunoreactivity decreased co-incidentally with the increase in PGP-9.5 immunoreactivityin the upper dermis (yellow). E-H) ProNGF and p75 association in sham-operated controls and in lesioned animals was loose in that most proNGF immunoreactivity was segregated from that for p75 and there was no obvious co-localization (arrows).

Article Snippet: Immunohistochemical staining with this antibody in either rat or human skin was absent following preadsorption with purified human PGP 9.5 protein [ , ]. proNGF: The rabbit anti-proNGF antibody (Alomone #ANT-005 Lot#AN-03) was generated by injection of a synthetic peptide corresponding to a.a. 84-104 of the precursor form of rat NGF.

Techniques: Staining, Immunostaining

Journal: Molecular Pain

Article Title: Neurotrophic factor changes in the rat thick skin following chronic constriction injury of the sciatic nerve

doi: 10.1186/1744-8069-8-1

Figure Lengend Snippet: Distribution of proNGF immunoreactivity in naïve glabrous skin and following application of CCI . A) In sham operated controls, proNGF (brown precipitate) was found in the upper dermis, particularly in wall of blood vessels, and in small patches of keratinocytes (arrowhead). B) 1 week post-injury, proNGF immunolabeling was detected in mast cells (arrows; identified based on toluidine blue metachromatic counterstaining), in blood vessels in the upper dermis as well as in keratinocytes. C) At 2 weeks post-injury, note the intense immunostaining of keratinocytes, the labeling of some unidentified structures in dermis and a lower number of immunostained mast cells. D) At 4 weeks post-injury, note mast cell immunolabeling. E) Preincubation of antibody with control peptide abolished all specific staining. F) proNGF quantification was done by Western Blot analysis (n = 6) from sham, and animals 1 - 4 week post-injury. OD was normalized against β-actin loading controls. ***p < 0.001, **p < 0.01, *p < 0.05; +SEM.

Article Snippet: Primary antibodies such as mouse anti-TrkA (1:500, MAB1056, R&D Systems, Minneapolis, MN), rabbit anti-proNGF (1:500, ANT-005, Alomone Labs, Isreal), or mouse anti-p75 (1:200, MC192, Novus Biologicals) were incubated overnight in blocking buffer at 4°C.

Techniques: Immunolabeling, Immunostaining, Labeling, Staining, Western Blot

Journal: Molecular Pain

Article Title: Neurotrophic factor changes in the rat thick skin following chronic constriction injury of the sciatic nerve

doi: 10.1186/1744-8069-8-1

Figure Lengend Snippet: Changes in p75 expression on S100-IR Schwann cells . A) In sham-operated rats Schwann cells (green) were detected by means of S100 immunoreactivity in small nerves, with immunoreactivity of varying intensity from bright (arrow), lower in the dermis, to dim, along the dermo-epidermal junction. P75 immunoreactivity (red) was evident surrounding all S100-IR Schwann cells. B) One week following nerve injury, p75 immunostaining was dramatically upregulated in Schwann cells; the intense red color masked the mixture of red and green stainings which could be detected by analysing the separately the S100 and p75 stainings (not shown). C-D) 2 & 4 weeks post-injury a decrease in p75 intensity was observed in that the yellow indicative of S100 co-labelling was able to be visualized. E) proNGF (red) S100 (green) and p75 (blue) triple labelling to demonstrate the relative distribution of Schwann cells with respect to proNGF; note a limited distribution of Schwann cells with proNGF and faint immunoreactivity for p75 in sham-operated controls. F) 2 weeks post-injury, the clear upregulation of p75 immunoreactivity associated with S100 (arrow) was observed, which wrapped around proNGF-IR blood vessels.

Article Snippet: Primary antibodies such as mouse anti-TrkA (1:500, MAB1056, R&D Systems, Minneapolis, MN), rabbit anti-proNGF (1:500, ANT-005, Alomone Labs, Isreal), or mouse anti-p75 (1:200, MC192, Novus Biologicals) were incubated overnight in blocking buffer at 4°C.

Techniques: Expressing, Immunostaining

Journal: Molecular Pain

Article Title: Neurotrophic factor changes in the rat thick skin following chronic constriction injury of the sciatic nerve

doi: 10.1186/1744-8069-8-1

Figure Lengend Snippet: Distribution of p75 immunoreactivity following nerve injury and relationship to nerve fibers and proNGF . A) In sham-operated animals, p75 was distributed (red) around and with PGP 9.5-IR nerve fibers (green). P75 staining was found more clearly around large cutaneous PGP 9.5-IR nerve fiber bundles and smaller fibers along the dermo-epidermal junction. Where nerve fibers crossed the dermo-epidermal junction into the epidermis, the yellow color representing p75 associating with nerve fibers was lost (arrow). B) At 1 week post-injury, virtually all PGP-9.5-IR nerve fibers disappeared from the upper dermis and epidermis; immunostaining in p75-IR Schwann was very intense. C) At 2 weeks post-injury, a low number of PGP-9.5-IR fibers were detected and were associated with p75 Schwann cells (yellow), however most of the PGP-9.5-IR was restricted to Langerhans cells in epidermis (arrow) D) At 4 weeks post-injury, p75 immunoreactivity decreased co-incidentally with the increase in PGP-9.5 immunoreactivityin the upper dermis (yellow). E-H) ProNGF and p75 association in sham-operated controls and in lesioned animals was loose in that most proNGF immunoreactivity was segregated from that for p75 and there was no obvious co-localization (arrows).

Article Snippet: Primary antibodies such as mouse anti-TrkA (1:500, MAB1056, R&D Systems, Minneapolis, MN), rabbit anti-proNGF (1:500, ANT-005, Alomone Labs, Isreal), or mouse anti-p75 (1:200, MC192, Novus Biologicals) were incubated overnight in blocking buffer at 4°C.

Techniques: Staining, Immunostaining

Journal: Molecular Pain

Article Title: Neurotrophic factor changes in the rat thick skin following chronic constriction injury of the sciatic nerve

doi: 10.1186/1744-8069-8-1

Figure Lengend Snippet: Distribution of p75 immunoreactivity following nerve injury and relationship to nerve fibers and proNGF . A) In sham-operated animals, p75 was distributed (red) around and with PGP 9.5-IR nerve fibers (green). P75 staining was found more clearly around large cutaneous PGP 9.5-IR nerve fiber bundles and smaller fibers along the dermo-epidermal junction. Where nerve fibers crossed the dermo-epidermal junction into the epidermis, the yellow color representing p75 associating with nerve fibers was lost (arrow). B) At 1 week post-injury, virtually all PGP-9.5-IR nerve fibers disappeared from the upper dermis and epidermis; immunostaining in p75-IR Schwann was very intense. C) At 2 weeks post-injury, a low number of PGP-9.5-IR fibers were detected and were associated with p75 Schwann cells (yellow), however most of the PGP-9.5-IR was restricted to Langerhans cells in epidermis (arrow) D) At 4 weeks post-injury, p75 immunoreactivity decreased co-incidentally with the increase in PGP-9.5 immunoreactivityin the upper dermis (yellow). E-H) ProNGF and p75 association in sham-operated controls and in lesioned animals was loose in that most proNGF immunoreactivity was segregated from that for p75 and there was no obvious co-localization (arrows).

Article Snippet: Immunohistochemical staining with this antibody in either rat or human skin was absent following preadsorption with purified human PGP 9.5 protein [ , ]. proNGF: The rabbit anti-proNGF antibody (Alomone #ANT-005 Lot#AN-03) was generated by injection of a synthetic peptide corresponding to a.a. 84-104 of the precursor form of rat NGF.

Techniques: Staining, Immunostaining

Journal: Molecular Vision

Article Title: Electroporation-mediated gene delivery of cleavage-resistant pro–nerve growth factor causes retinal neuro- and vascular degeneration

doi:

Figure Lengend Snippet: Overexpression of proNGF reduced NGF and induced expression of TrkA and p75 NTR , but not sortilin. A , B : Western blot (WB) analysis of rat retinal lysate showed significant (1.5-fold) increase in expression of TrkA and p75 NTR in rats electroporated with pGFP-proNGF123 as compared with those electroporated with pGFP (n=4). C : WB analysis showed that overexpression of pGFP-proNGF123 in rat retinas did not affect sortilin protein expression level as compared with the control rats (n=4). D : WB analysis showed that overexpression of pGFP-proNGF123 significantly reduced NGF expression (n=5) compared with GFP controls. The asterisk represents significant difference as compared with control group at p<0.05. ROD, relative optical density.

Article Snippet: Antibodies for proNGF and NGF (Alomone Labs Ltd), p75 NTR , sortilin, and TrkA (Millipore) were used.

Techniques: Over Expression, Expressing, Western Blot