The pathogenesis of type 2 diabetes mellitus (T2DM) is commonly associated with altered gut bacteria. However, whether the microbial dysbiosis that exists in human diabetic patients with or without retinopathy is different remains largely unknown. Here, we collected clinical information and fecal samples from 75 participants, including 25 diabetic patients without retinopathy (DM), 25 diabetic patients with retinopathy (DR), and 25 healthy controls (HC). The gut microbial composition in the three groups was analyzed using 16S ribosomal RNA (rRNA) gene sequencing. Microbial structure and composition differed in the three groups. The α and β diversities in both the DM and DR groups were reduced compared with those in the HC group. Blautia was the most abundant genus, especially in the DM group. In addition, increased levels of Bifidobacterium and Lactobacillus and decreased levels of Escherichia-Shigella, Faecalibacterium, Eubacterium_hallii_group and Clostridium genera were observed in the DM and DR groups compared with the HC group. Furthermore, a biomarker set of 25 bacterial families, which could distinguish patients in the DR group from those in the DM and HC groups was identified, with the area under the curve values ranging from 0.69 to 0.85. Of note, Pasteurellaceae, which was increased in DM and decreased in DR compared with HC, generated a high AUC (0.74) as an individual predictive biomarker. Moreover, 14 family biomarkers were associated with fasting blood glucose levels or diabetes, with most of them being negatively correlated. In summary, our study establishes compositional alterations of gut microbiota in DM and DR, suggesting the potential use of gut microbiota as a non-invasive biomarker for clinical and differential diagnosis, as well as identifying potential therapeutic targets of diabetic retinopathy.
In this study we investigated the biocompatibility of collagen-chitosan-sodium hyaluronate (Col-Chi-NaHA) complexes and cornea tissue, and the feasibility of Col-Chi-NaHA complexes as substrates for cultivating rabbit corneal cells. Different components of Col-Chi-NaHA complexes were prepared and tested. A circular complex film with a diameter of 6 mm was inserted into rabbit stomal pocket and traced for a period of 5 months. Clinical examination was made. Rabbit limbal corneal epithelial cells, corneal endothelial cells, and keratocytes were cultured primarily on complexes. Phase contrast microscope examination was made daily. Histological, immunohistochemical, and scanning electron microscopic examinations were carried out. The complexes of 20% collagen, 10% chitosan, and 0.5% sodium hyaluronate showed rather weak corneal edema and other responses. The degradation of materials was obvious after 5 months. Corneas were transparent and translucent. Cells seeded on Col-Chi-NaHA were allowed to proliferate and partly form confluent monolayer after 9 days in culture. Cultured cells were well attached to the complexes of 20% collagen, 10% chitosan, and 0.5% sodium hyaluronate, or 10% chitosan and 0.5% sodium hyaluronate. The results showed that Col-Chi-NaHA complexes had good biocompatibility with cornea. The complexes can degrade and be absorbed in cornea. Col-Chi-NaHA complex may be a suitable substrate for cultivating corneal cells and a feasible material as a scaffold of tissue-engineered cornea.
The development of the cornea, a highly specialized transparent tissue located at the anterior of the eye, is coordinated by a variety of molecules and cells. Here, we report that mast cells (MCs), recently found to be involved in morphogenesis, played a potentially important role in corneal development in mice. We show that two different waves of MC migration occurred during corneal development. In the first wave, MCs migrated to the corneal stroma and became distributed throughout the cornea. This wave occurred by embryonic day 12.5, with MCs disappearing from the cornea at the time of eyelid opening. In the second wave, MCs migrated to the corneal limbus and became distributed around limbal blood vessels. The number of MCs in this region gradually increased after birth and peaked at the time of eyelid opening in mice, remaining stable after postnatal day 21. We also show that integrin α4β7 and CXCR2 were important for the migration of MC precursors to the corneal limbus and that c-Kit-dependent MCs appeared to be involved in the formation of limbal blood vessels and corneal nerve fibers. These data clearly revealed that MCs participate in the development of the murine cornea.
Type I collagen membranes with tailored fibril nanoarchitectures were fabricated through a vitrification processing, which mimicked, to a degree, the collagen maturation process of corneal stromal extracellular matrix in vivo. Vitrification was performed at a controlled temperature of either 5 °C or 39 °C at a constant relative humidity of 40% for various time periods from 0.5 wk up to 8 wk. During vitrification, the vitrified collagen membranes (collagen vitrigels, CVs) exhibited a rapid growth in fibrillar density through the evaporation of water and an increase in fibrillar stiffness due to the formation of new and/or more-stable interactions. On the other hand, the collagen fibrils in CVs maintained their D-periodicity and showed no significant difference in fibrillar diameter, indicating preservation of the native states of the collagen fibrils during vitrification. Keratocyte phenotype was maintained on CVs to varying degrees that were strongly influenced by the collagen fibril nanoarchitectures. Specifically, the vitrification time of CVs mainly governed the keratocyte morphology, showing significant increases in the cell protrusion number, protrusion length, and cell size along with CV vitrification time. The CV vitrification temperature affected the regulation of keratocyte fibroblasts' gene expressions, including keratocan and aldehyde dehydrogenase (ALDH), demonstrating a unique way to control the expression of specific genes in vitro.
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