Browsing by Author "Dominguez, James M."

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    Bone Marrow–Derived Cell Recruitment to the Neurosensory Retina and Retinal Pigment Epithelial Cell Layer Following Subthreshold Retinal Phototherapy
    (ARVO, 2017-10) Caballero, Sergio; Kent, David L.; Sengupta, Nilanjana; Li Calzi, Sergio; Shaw, Lynn; Beli, Eleni; Moldovan, Leni; Dominguez, James M.; Moorthy, Ramana S.; Grant, Maria B.; Medicine, School of Medicine
    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.
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    Combination therapies prevent the neuropathic, proinflammatory characteristics of bone marrow in streptozotocin-induced diabetic rats
    (American Diabetes Association, 2015-02) Dominguez, James M.; Yorek, Mark A.; Grant, Maria B.; Department of Ophthalmology, IU School of Medicine
    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-1β, 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.
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    CX3CR1 deficiency accelerates the development of retinopathy in a rodent model of type 1 diabetes
    (Springer, 2016-11) Beli, Eleni; Dominguez, James M.; Hu, Ping; Thinschmidt, Jeffrey S.; Caballero, Sergio; Calzi, Sergio Li; Luo, Defang; Shanmugam, Sumathi; Salazar, Tatiana; Duan, Yaqian; Boulton, Michael E.; Mohr, Susanna; Abcouwer, Steven F.; Saban, Daniel R.; Harrison, Jeffrey K.; Grant, Maria B.; Ophthalmology, School of Medicine
    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. 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.
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    Tumor Necrosis Factor Alpha (TNF-α) Disrupts Kir4.1 Channel Expression Resulting in Müller Cell Dysfunction in the RetinaDiurnal Rhythm of Kir4.1 in the Retina
    (ARVO, 2017-05-01) Hassan, Iraj; Luo, Qianyi; Majumdar, Sreeparna; Dominguez, James M.; Busik, Julia V.; Bhatwadekar, Ashay D.; Department of Ophthalmology, IU School of Medicine
    Purpose: Diabetic patients often are affected by vision problems. We previously identified diabetic retinopathy (DR) as a disease of clock gene dysregulation. TNF-α, a proinflammatory cytokine, is known to be elevated in DR. Müller cells maintain retinal water homeostasis and K+ concentration via Kir4.1 channels. Notably, Kir4.1 expression is reduced in diabetes; however, the interplay of TNF-α, Kir4.1, and clock genes in Müller cells remains unknown. We hypothesize that the Kir4.1 in Müller cells is under clock regulation, and increase in TNF-α is detrimental to Kir4.1. Methods: Long-Evans rats were made diabetic using streptozotocin (STZ). Retinal Kir4.1 expression was determined at different time intervals. Rat Müller (rMC-1) cells were transfected with siRNA for Per2 or Bmal1 and in parallel treated with TNF-α (5–5000 pM) to determine Kir4.1 expression. Results: Kir4.1 expression exhibited a diurnal rhythm in the retina; however, with STZ-induced diabetes, Kir4.1 was reduced overall. Kir4.1 rhythm was maintained in vitro in clock synchronized rMC-1 cells. Clock gene siRNA-treated rMC-1 exhibited a decrease in Kir4.1 expression. TNF-α treatment of rMCs lead to a profound decrease in Kir4.1 due to reduced colocalization of Kir4.1 channels with synapse-associated protein (SAP97) and disorganization of the actin cytoskeleton. Conclusions: Our findings demonstrate that Kir4.1 channels possess a diurnal rhythm, and this rhythm is dampened with diabetes, thereby suggesting that the increase in TNF-α is detrimental to normal Kir4.1 rhythm and expression.