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. 2018 Jan-Dec:14:1744806917751322.
doi: 10.1177/1744806917751322.

CCR2 upregulation in DRG neurons plays a crucial role in gastric hyperalgesia associated with diabetic gastropathy

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CCR2 upregulation in DRG neurons plays a crucial role in gastric hyperalgesia associated with diabetic gastropathy

Aye Aye-Mon et al. Mol Pain. 2018 Jan-Dec.

Abstract

Background Diabetic gastropathy is a complex neuromuscular dysfunction of the stomach that commonly occurs in diabetes mellitus. Diabetic patients often present with upper gastrointestinal symptoms, such as epigastric discomfort or pain. The aim of this study was to assess gastric sensation in streptozocin-induced diabetes mellitus (DM) rats and to determine the contribution of C-C motif chemokine receptor 2 (CCR2) signaling to gastric hyperalgesia. Results DM rats showed signs of neuropathy (cutaneous mechanical hyperalgesia) from two weeks after streptozocin administration until the end of the experiment. Accelerated solid gastric emptying was observed at two weeks after streptozocin administration compared to the controls. Intense gastric hyperalgesia also developed in DM rats at two weeks after streptozocin administration, which was significantly reduced after intrathecal administration of the CCR2 antagonist INCB3344. Immunochemical analysis indicated that CCR2 expression was substantially upregulated in small and medium-sized dorsal root ganglia neurons of DM rats, although the protein level of monocyte chemoattractant protein-1, the preferred ligand for CCR2, was not significantly different between the control and DM groups. Conclusions These data suggest that CCR2 activation in nociceptive dorsal root ganglia neurons plays a role in the pathogenesis of gastric hyperalgesia associated with diabetic gastropathy and that CCR2 antagonist may be a promising treatment for therapeutic intervention.

Keywords: CCR2 receptor; Neuropathic pain; diabetes mellitus; gastric hyperalgesia; streptozocin.

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Figures

Figure 1.
Figure 1.
Differences in body weight and blood glucose level between the control and diabetic groups. Body weight (a) and random blood glucose level (b) were measured in the control (CT) and diabetic (DM) rats. Data are shown as mean ± SEM, *p < 0.001 CT versus DM, n = 15–20 rats per group. No weight gain (a) and hyperglycemia (b) were observed in DM rats throughout the experiments.
Figure 2.
Figure 2.
Changes in cutaneous sensitivities after induction of diabetes by STZ. The Randall-Selitto test (a), von Frey’s test (b), and heat sensitivity test (c) were performed in control (CT) and diabetic (DM) rats. Data are shown as mean ± SEM, *p < 0.001, p < 0.05 CT versus DM, n = 6 rats per group. PWT measured by Randall-Selitto test (a) and von Frey’s test (b) was reduced in the STZ-induced DM group.
Figure 3.
Figure 3.
Effect of STZ-induced diabetes on gastric motor and sensory functions. Gastric emptying (a) and gastric sensitivity (b) were assessed in control (CT) and diabetic (DM) rats. Data are shown as mean ± SEM, *p < 0.001, #p < 0.01 CT versus DM. (a) Significantly rapid gastric emptying was observed in DM rats at two weeks after STZ administration (n = 6 rats per group). (b) Distension-induced gastric hyperalgesia developed significantly in DM rats (n = 9 rats per group). (c) Representative EMG recording data from each CT and DM rats.
Figure 4.
Figure 4.
Effect of CCR2 antagonist (INCB3344) on diabetes-induced gastric hyperalgesia. Gastric sensitivity was measured in the control (CT) and diabetic (DM) rats at two weeks after STZ administration. The CCR2 antagonist INCB3344 (CCR2-A) or vehicle (veh) was injected intrathecally. Data are shown as mean ± SEM, #p < 0.01, p < 0.05 veh versus CCR2-A, n = 5–7 rats per group. INCB3344 (1 mM and 0.1 mM) significantly suppressed distension-induced gastric hyperalgesia in DM rats at 1 and 2 h after administration as compared with vehicle-treated rats.
Figure 5.
Figure 5.
Cellular localization of CCR2-IR in T10 DRG at two weeks after STZ administration. Representative images of immunostaining with anti-CCR2 antibody in control (CT) (a) and diabetic (DM) (b) rats. Arrows represent CCR2-positive cells. Scale bar: 50 µm. Number of total and CCR2-IR neurons per section (c, e) and their size distribution (d, f) were determined in both the CT and DM groups. Data are shown as mean ± SEM, *p < 0.001 CT versus DM, n = 4 rats per group. CCR2 was exclusively expressed in small- and medium-sized DRG neurons in both the CT and DM rats (a, b, f). CCR2-IR neurons were significantly more numerous in the DM group than in the CT (a, b, e) despite no significant change between them in total DRG neurons per section (c). The number of CCR2-IR cells was relatively greater in small DRG neurons of DM rats according to the size distribution of the DRG neurons (d, f).
Figure 6.
Figure 6.
CCR2 protein expression in T10 spinal cord at two weeks after STZ administration. Representative images of immunostaining with anti-CCR2 antibody in control (CT) (a) and diabetic (DM) (b) rats. Arrows represent CCR2-positive regions. (a2) and (b2) are higher magnification images of white-boxed regions in (a1) and (b1), respectively. Scale bar: 50 µm. CCR2-IR was detected only in the superficial layer of the spinal dorsal horn (n = 4 rats per group). (c) Representative Western blot analyses using anti-CCR2 and GAPDH antibodies. GAPDH was used as a loading control. CT 1–3: CT rats; DM 1–3: DM rats. (d) Semi-quantitative densitometric data for CCR2-IR are shown. Data are shown as mean ± SEM, n = 3 rats per group. The CCR2 protein level in the spinal cord was not significantly different between the CT and DM groups.
Figure 7.
Figure 7.
Immunolocalization of MCP-1 and its protein level in T10 DRG at two weeks after STZ administration. Representative images of immunostaining with anti-MCP-1 antibody in control (CT) (a) and diabetic (DM) (b) rats. Scale bar: 50 µm. MCP-1 was constitutively expressed in DRG neurons in both CT and DM rats (n = 4 rats per group). (c) Representative Western blot analyses using anti-MCP-1 and GAPDH antibodies are shown. GAPDH was used as a loading control. CT 1–3: CT rats; DM 1–3: DM rats. (d) Semi-quantitative densitometric data for MCP-1-IR is shown. Data are shown as mean ± SEM, n = 3 rats per group. There was no difference in MCP-1 protein levels between the CT and DM groups.
Figure 8.
Figure 8.
Immunolocalization of MCP-1 and its protein level in T10 spinal cord at two weeks after STZ administration. Representative images of immunostaining with anti-MCP-1 antibody in control (CT) (a) and diabetic (DM) (b) rats. (a2) and (b2) are higher magnification images of white-boxed regions in (a1) and (b1), respectively. Scale bar: 50 µm. MCP-1 was expressed in the whole spinal cord (n = 4 rats per group). Double-labeling IHC showed MCP-1 to be co-localized with astrocyte marker (GFAP) (c), but not with microglial (Iba1) (d) and neuronal (NeuN) (e) markers. Scale bar: 50 µm. (f) Representative Western blot analyses using anti-MCP-1 and GAPDH antibodies are shown. GAPDH was used as a loading control. CT 1–3: CT rats; DM 1–3: DM rats. (g) Semi-quantitative densitometric data for MCP-1-IR are shown. Data are shown as mean ± SEM, n = 3 rats per group. The MCP-1 protein level was higher in the DM group than in the CT group, but the data were not significant.

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References

    1. International Diabetes Federation. IDF Diabetes Atlas, 7th ed. 2015, https://www.idf.org/e-library/epidemiology-research/diabetes-atlas.html
    1. Boulton AJ, Vinik AI, Arezzo JC, et al. Diabetic neuropathies: a statement by the American Diabetes Association. Diabetes Care 2005; 28: 956–962. - PubMed
    1. Feldman EL. Epidemiology and classification of diabetic neuropathy – up to date, https://www.uptodate.com (2015, accessed 5 January 2017).
    1. Vinik AI, Maser RE, Mitchell BD, et al. Diabetic autonomic neuropathy. Diabetes Care 2003; 26: 1553–1579. - PubMed
    1. Bharucha AE, Camilleri M, Forstrom L, et al. Relationship between clinical features and gastric emptying disturbances in diabetes mellitus. Clin Endocrinol 2009; 70: 415–420. - PMC - PubMed

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