Bone Marrow-Derived Cell Recruitment to the Neurosensory Retina and Retinal Pigment Epithelial Cell Layer Following Subthreshold Retinal Phototherapy.

Purpose: We investigated whether subthreshold retinal phototherapy (SRPT) was associated with recruitment of bone marrow (BM)-derived cells to the neurosensory retina (NSR) and RPE layer. Methods: GFP chimeric mice and wild-type (WT) mice were subjected to SRPT using a slit-lamp infrared laser. Duty cycles of 5%, 10%, 15%, and 20% (0.1 seconds, 250 mW, spot size 50 µm) with 30 applications were placed 50 to 100 µm from the optic disc. In adoptive transfer studies, GFP+ cells were given intravenously immediately after WT mice received SRPT. Immunohistochemistry was done for ionized calcium-binding adapter molecule-1 (IBA-1+), CD45, Griffonia simplicifolia lectin isolectin B4, GFP or cytokeratin). Expression of Ccl2, Il1b, Il6, Hspa1a, Hsp90aa1, Cryab, Hif1a, Cxcl12, and Cxcr4 mRNA and flow cytometry of the NSR and RPE-choroid were performed. Results: Within 12 to 24 hours of SRPT, monocytes were detected in the NSR and RPE-choroid. Detection of reparative progenitors in the RPE occurred at 2 weeks using flow cytometry. Recruitment of GFP+ cells to the RPE layer occurred in a duty cycle-dependent manner in chimeric mice and in mice undergoing adoptive transfer. Hspa1a, Hsp90aa1, and Cryab mRNAs increased in the NSR at 2 hours post laser; Hif1a, Cxcl12, Hspa1a increased at 4 hours in the RPE-choroid; and Ccl2, Il1b, Ifng, and Il6 increased at 12 to 24 hours in the RPE-choroid. Conclusions: SRPT induces monocyte recruitment to the RPE followed by hematopoietic progenitor cell homing at 2 weeks. Recruitment occurs in a duty cycle-dependent manner and potentially could contribute to the therapeutic efficacy of SRPT.

CX3CR1 deficiency accelerates the development of retinopathy in a rodent model of type 1 diabetes.

In this study, the role of CX3CR1 in the progression of diabetic retinopathy (DR) was investigated. The retinas of wild-type (WT), CX3CR1 null (CX3CR1gfp/gfp, KO), and heterozygous (CX3CR1+/gfp, Het) mice were compared in the presence and absence of streptozotocin (STZ)-induced diabetes. CX3CR1 deficiency in STZ-KO increased vascular pathology at 4 months of diabetes, as a significant increase in acellular capillaries was observed only in the STZ-KO group. CX3CR1 deficiency and diabetes had similar effects on retinal neurodegeneration measured by an increase in DNA fragmentation. Retinal vascular pathology in STZ-KO mice was associated with increased numbers of monocyte-derived macrophages in the retina. Furthermore, compared to STZ-WT, STZ-KO mice exhibited increased numbers of inflammatory monocytes in the bone marrow and impaired homing of monocytes to the spleen. The induction of retinal IL-10 expression by diabetes was significantly less in KO mice, and when bone marrow-derived macrophages from KO mice were maintained in high glucose, they expressed significantly less IL-10 and more TNF-α in response to LPS stimulation. These findings support that CX3CR1 deficiency accelerates the development of vascular pathology in DR through increased recruitment of proinflammatory myeloid cells that demonstrate reduced expression of anti-inflammatory IL-10. KEY MESSAGES: • CX3CR1 deletion in STZ-diabetic mice accelerated the onset of diabetic retinopathy (DR). • The early onset of DR was associated with increased retinal cell apoptosis. • The early onset of DR was associated with increased recruitment of bone marrow-derived macrophages to the retina. • Bone marrow-derived macrophages from CX3CR1 KO diabetic mice expressed more TNF-α and less IL-10. • The role of IL-10 in protection from progression of DR is highlighted.

Combination therapies prevent the neuropathic, proinflammatory characteristics of bone marrow in streptozotocin-induced diabetic rats.

We previously showed that peripheral neuropathy of the bone marrow was associated with loss of circadian rhythmicity of stem/progenitor cell release into the circulation. Bone marrow neuropathy results in dramatic changes in hematopoiesis that lead to microvascular complications, inflammation, and reduced endothelial repair. This series of events represents early pathogenesis before development of diabetic retinopathy. In this study we characterized early alterations within the bone marrow of streptozotocin (STZ)-induced diabetic rats following treatments that prevent experimental peripheral neuropathy. We asked whether bone marrow neuropathy and the associated bone marrow pathology were reversed with treatments that prevent peripheral neuropathy. Three strategies were tested: inhibition of neutral endopeptidase, inhibition of aldose reductase plus lipoic acid supplementation, and insulin therapy with antioxidants. All strategies prevented loss of nerve conduction velocity resulting from STZ-induced diabetes and corrected the STZ-induced diabetes-associated increase of immunoreactivity of neuropeptide Y, tyrosine hydroxylase, and somatostatin. The treatments also reduced concentrations of interleukin-1ß, granulocyte colony-stimulating factor, and matrix metalloproteinase 2 in STZ-induced diabetic bone marrow supernatant and decreased the expression of NADPH oxidase 2, nitric oxide synthase 2, and nuclear factor-κB1 mRNA in bone marrow progenitor cells. These therapies represent novel approaches to attenuate the diabetic phenotype within the bone marrow and may constitute an important therapeutic strategy for diabetic microvascular complications.