Epigenetic modifications such as DNA methylation, histone post-translational modifications, non-coding RNA are reversible, heritable alterations which are induced by environmental stimuli. Major risk factors of diabetes and diabetic complications including hyperglycemia, oxidative stress and advanced glycation end products, can lead to abnormal epigenetic modifications in retinal vascular endothelial cells and retinal pigment epithelium cells. Epigenetic mechanisms are involved in the pathogenesis of macular edema and neovascularization of diabetic retinopathy (DR), as well as diabetic metabolic memory. The heritable nature of epigenetic marks also playsakey role in familial diabetes mellitus. Further elucidation of epigenetic mechanisms in DR can open the way for the discovery of novel therapeutic targets to prevent DR progression.
In thiis study,we show thai carbachol stimulates the accumulation of inositol phosphates(InsPs)in human rellnal pigment epithelium (RPE)cells and atropine blocks the carbachol-induced effect ,suggesting the existence of musearinie acelyleholine receptors in human RPE cells. In contrast,noradrenaline,serotonin, cpidermal growth factor (EGF),isoproterenol,and NECA (5'-[N-ethyl]-carboxamido-adenosine)do not influence the basal levels of InsPs.Moreover,isoprmerenol and NECA do not affect the carhaehol elevated levels of InsPs.EGF,howcvcr,does potentiate the carhaehol stimulated elevation of InsPs in a dose-dependent manner ,suggesting an interaction between EGF and musearinie receptors in cultured human RPE cells. （Chin J Ocul Fundus Dis,1994,10:220-222）
The ultra.structural changes of photic injury to the retina of two patients caused by indirect ophthalmoscope were studied.The duration of exposure was 20 minutes. Under transmission electron microscope, dilation of the choroidal vessels,swelling and vacule formation at all retinal layers, disparsed pyknotic nuclei among the outer nuclear layer and swelling of the nerve fibers were found,The retina of the pregnant woman was injured more severely than that of the male patient. （Chin J Ocul Fundus Dis,1994,10:77-79）
Objective To observe the expression of cyclin dependent kinase 5 (Cdk5) and p25 in the pathogenesis of retinitis pigmentosa (RP) in Royal College of Surgeon (RCS) rats and its relationships with apoptosis. To explore the mechanism of Cdk5 and p25 induced photoreceptor apoptosis in the pathogenesis of RP. Methods Retinas of RCS and RCS-rdy+ rats were obtained at the ages of postnatal day 17, 25, 35, 60. The retinal structure and thickness of outer nuclear layer were measured by optical microscopy. The expression of Cdk5, p25, cleave-caspase 3 in the retina was evaluated by immunohistochemistry. The protein expression of cleave-caspase 3 in the retina was determined by Western blot. The apoptosis of retinal cells was examined by terminal-deoxynucleotidyl transferase mediated nick end labeling (TUNEL). The mean absorbance value of apoptotic cells was analyzed by SPSS 17.0 software. Results The retinal thickness of the RCS rats was significantly reduced in comparison to the RCS-rdy+ rats as the postnatal days progressed, particularly in the layer of rods and cones and the outer nuclear layer. The expression level of Cdk5, p25, cleave-caspase 3 of RCS rats increased from postnatal 17 days to postnatal 35 days, while decreased on postnatal 60 days; but there was no obvious change of above indexes in RCS-rdy+ rats. The protein expression of cleave-caspase 3 in the RCS rats was significantly increased with progression of postnatal days to postnatal 35; but there was no obvious similar change in RCS-rdy+ rats. The results of TUNEL showed that the apoptotic cells significantly increased in the outer nuclear layer of RCS rats from postnatal 17 days to postnatal 35 days, while decreased on postnatal 60 days; but there was no obvious change of above index in RCS-rdy+ rats. This study showed that there were significant correlations between the following variables: Cdk5 expression and p25 expression, Cdk5 expression and cleave-caspase 3 expression, Cdk5 expression and apoptotic cells, p25 expression and cleave-caspase 3 expression, p25 expression and apoptotic cells, cleave-caspase 3 expression and apoptotic cells. The partial correlation coefficients were 0.949, 0.808, 0.959, 0.887, 0.979, 0.852, respectively and the P value was 0.000. Conclusions The apoptotic cells significantly increases and the expression level of Cdk5, p25, cleave-caspase 3 of RCS rats increases from postnatal 17 days to postnatal 35 days. The tendency of apoptotic cells to increase is consistent with the change of Cdk5, p25, cleave-caspase 3 expression. The apoptosis of photoreceptor cells is related to increasing expression of Cdk5 and p25 in RCS rats. Cdk5 may be involved in the development of RP in RCS rats.
OBJECTIVE: To isolate and characterize mesenchymal stem cells (MSCs) derived from bone marrow of Banna minipig inbred line (BMI). METHODS: BMI-MSCs was isolated from bone marrow by density gradient centrifugation and cultured in DMEM (containing 15% bovine serum) at 37 degrees C with humidified 5% CO2. These cultured stem cells were characterized in clonal growth, expression of specific markers and capability of differentiation. RESULTS: Mesenchymal stem cells were proliferative and could be expanded rapidly in vitro. Clonal growth of these cells can be observed when small amount of cells was inoculated. These cells were SH2, SH3, SH4, SB10 and SB21 positive. And it was proved that these cells possess osteo-differentiation ability, up-regulated alkaline phosphatase expression and calcium secretion after osteosupplement was added into the media for several days. CONCLUSION: Mesenchymal stem cells derived from bone marrow of BMI possess the general characters of stem cell.
Objective To investigate the effectiveness of free superficial circumflex iliac artery flap (SCIP) combined with deep inferior epigastric perforator flap (DIEP) in repairing large soft tissue defects of upper extremities. MethodsBetween February 2017 and January 2021, free SCIP combined with DIEP was used to repair 15 patients with large soft tissue defects of upper extremities. There were 12 males and 3 females, aged from 34 to 52 years, with a median age of 41 years. The causes of injury were mechanical injury in 11 cases and traffic accident injury in 4 cases. There were 9 cases of circular skin defect in the forearm and 6 cases of skin defect around the upper arm and elbow joint, with the defect in size of 15.5 cm×10.5 cm to 26.5 cm×15.5 cm, accompanied by exposure of deep tissues such as tendons and bones. There were 7 cases with open fractures and 8 cases with vascular and nerve injuries. The time from injury to operation ranged from 7 to 14 days, with an average of 7.5 days. The flaps in size of 16.0 cm×11.0 cm to 27.0 cm×16.0 cm were harvested; the thickness of the flaps was 0.8-1.3 cm, and the excess fat tissue was removed under the microscope after harvesting. The length of proximal vascular pedicle was 5.0-7.0 cm, and of distal vascular pedicle was 3.0-5.0 cm. The donor site was closed and sutured directly, and the navel was reconstructed. Results The flaps survived successfully in 14 cases, and the arterial crisis occurred in 1 case at 10 hours after operation, and the flap survived after surgical exploration. All the wounds in the donor and recipient sites healed by first intention. All 15 patients were followed up 3-14 months, with an average of 10.5 months. The appearance, elasticity, and texture of flap were good without obvious bloat, contracture, or adhesion. The donor site healed well, no abdominal wall hernia was found, and the appearance of reconstructed navel was good, only linear scar left. At last follow-up, 12 cases were evaluated as excellent and 3 cases were good according to the evaluation criteria of flap function of the Chinese Medical Association Microsurgery Branch. The sensation recovered to \begin{document}${\rm{S}}_{3^+} $\end{document} in 7 cases and \begin{document}${\rm{S}}_3 $\end{document} in 8 cases. ConclusionFree SCIP combined with DIEP has a large excision area and excellent skin ductility. It is an effective clinical method for repairing large soft tissue defects of upper extremities.
Waardenburg syndrome is a rare genetic disease of auditory pigmentation. The main symptom is sensorineural hearing loss. Pigment disorders and other developmental defects in skin, hair, iris, fundus and other parts are specifically divided into four different subtypes, each of which corresponds to different pathogenic genes, which encode transcription factors and signaling molecules that play a key role in the development process of neural crest cells into melanocytes. Because there are multiple subtypes of Waardenburg syndrome, different subtypes exhibit different symptoms, signs and ocular manifestations. Patients with Waardenburg syndrome are often first treated in ENT head and neck surgery due to hearing loss. Lack of theoretical knowledge related to Waardenburg syndrome by ophthalmologists may lead to misdiagnosis or missed diagnosis. Although there are currently limited treatments for the disease, with the continuous development of gene therapy and hearing management methods, the future treatment prospects will be broader.
Objective To summarize the anatomical types of the concomitant veins of deep inferior epigastric artery and the corresponding choice of anastomosis methods, and to evaluate the indications and safety of various methods. MethodsBetween October 2015 and June 2021, 296 female patients received breast reconstruction with autologous free lower abdominal flap, including 154 cases of immediate breast reconstruction and 142 cases of delayed breast reconstruction. The average age of the patients was 36.5 years, ranged from 26 to 62 years. Unilateral free deep inferior epigastric artery perforator flap transplantation was performed in 172 cases, and unilateral free muscle-sparing rectus abdominis myocutaneous flap transplantation was performed in 124 cases. The internal thoracic vessels were selected as the recipient vessels in all cases. The length of vascular pedicle ranged from 9.5 to 13.0 cm, with an average of 11.2 cm. The concomitant veins of deep inferior epigastric artery included three anatomical types: one-branch type in 26 cases (8.8%), two-branch type in 180 cases (60.8%), and Y-shaped structure in 90 cases (30.4%). Anastomosis of inferior epigastric artery was performed with the proximal end of internal thoracic artery, while that of vein should be adjusted according to the diameter, length, wall thickness, and branches of the vein. The specific classification of venous anastomosis included: ① The sole concomitant vein of deep inferior epigastric artery was anastomosed with the internal mammary vein (26 cases); ② The two concomitant veins of deep inferior epigastric artery were anastomosed with the internal mammary vein respectively (151 cases); ③ The larger one of two concomitant veins of deep inferior epigastric artery was anastomosed with the internal mammary vein, and the other one was ligated and discarded (29 cases); ④ The two concomitant veins of deep inferior epigastric artery were Y-shaped, and the common trunk segment was directly anastomosed with the internal mammary vein (31 cases); ⑤ The smaller branch of the concomitant veins of Y-shaped structure was ligated, and the thicker branch was anastomosed with the internal mammary vein (17 cases); ⑥ The concomitant veins of Y-shaped structure were anastomosed with the internal mammary veins respectively (42 cases). Results Among the patients who underwent venous anastomosis according to scheme ④, 3 cases had venous crisis of flap after operation, of which 1 case was successfully rescued by emergency exploration; the other 2 cases had flap necrosis, of which 1 case survived after being repaired by pedicled latissimus dorsi flap, and 1 case closed the wound directly. Flaps with other venous anastomosis protocols survived completely. All 296 patients were followed up 12-70 months, with an average of 37.5 months. The reconstructed breast has good shape, good elasticity, and no skin flap contracture. The donor site of the flap only left linear scar, and the abdominal wall function was not affected. ConclusionThe method of direct anastomosis of concomitant veins of deep inferior epigastric artery with Y-shaped structure is relatively risky, and the vessels are prone to be twisted and compressed, leading to the occurrence of venous crisis. It can improve the safety of surgery if only one large vein is anastomosed or two veins are separated to anastomose respectively.
ObjectiveTo investigate the role of sonic hedgehog (Shh) signal transduction pathway in the expression of vascular endothelial growth factor (VEGF) under hypoxia in cultured human retinal pigment epithelial (hRPE) cells. MethodsARPE-19 were cultured and divided into normal ARPE-19 (Cont) and hypoxia group (100 μmol/L CoCl2 Cobalt Chloride +ARPE-19); hypoxia group was further divided into CoCl2 group, cyclopamine group (CYA) and dimethyl sulfoxide (DMSO) group. 20μmol/L cyclopamine was added to the CYA group 1 hour before hypoxia, 1‰DMSO was added into DMSO group at the same time. The hRPE cells were cultured under hypoxia for 4, 8, 12, 24 hours. The expression of Shh and VEGF were determined by Real-time fluorescent quantitate PCR (RT-PCR). The amount of VEGF in the hRPE-conditioned supernatant was measured using enzyme linked immunosorbent assay (ELISA) at 4, 8, 12, 24 hours, respectively. ResultsRT-PCR tests showed that the level of Shh and VEGF of hRPE was time dependently increased (Shh: F=45.260, P=0.001; VEGF: F=264.938, P=0.001). The level of Shh and VEGF of hRPE in the group treated with cyclopamine was decreased (P < 0.01). ELISA tests showed that the amount of VEGF in hRPE supernatant was significantly increased in time-dependent manner (F=3 156.676, P=0.001), and it was down-regulated by cyclopamine under hypoxia (P < 0.01). ConclusionShh signal transduction pathway could play a role in the VEGF expression induced by hypoxia in hRPE cells.
Objective To investigate the effects of exosomes from cultured human retinal pigment epithelium (ARPE-19) cells affected by oxidative stress on the proliferation and expression of vascular endothelial growth factor-A (VEGF-A) and Akt of ARPE-19 cells. Methods Culture ARPE-19 cells. The concentration of 2.5 μmol/L rotenone was selected to simulate oxidative stress and isolated ARPE-19-exosome. Exosomes were isolated by ExoQuick exosome precipitation solution. Transmission electron microscopy was used to identify the morphology of exosomes. Western blot was used to detect exosomes’ surface-specific maker protein CD63. ARPE-19 cells affected by oxidative stress were cultured with exosome as experimental group, normal ARPE-19 cells were cultured with exosome as control group. The cell proliferation was examined by methyl thiazolyl tetrazolium assay. Western blot and immunofluorescence assay were used to detect the expression levels of VEGF-A and Akt protein. Real-time quantitative polymerase chain reaction (RT-PCR) was used to detect the levels of VEGF-A mRNA and Akt mRNA. Results The diameter of normal ARPE-19-exosomes ranged from 50 to 150 nm. The isolated exosomes expressed CD63. AREP-19 cells were cultured with ARPE-19 (affected by rotenone)-exosome, the cell viability in experimental group was significantly reduced than in the control group. Green fluorescence was observed in the cytoplasm under fluorescence microscope. Compared with the control group, VEGF-A was up-regulated expressed and Akt was down-regulated expressed. Western blot results showed that, VEGF-A protein expression in the experimental group were higher than the control group. Akt protein expression in the experimental group were less than the control group. The difference was statically significant (t=3.822, 6.527;P＜0.05). RT-PCR results showed that VEGF-A mRNA expression levels was higher in the experimental group than the control group. Akt mRNA expression levels was lower in the experimental group than the control group. The difference was statically significant (t=8.805, ?7.823;P＜0.05). Conclusions Exosomes from ARPE-19 cells affected by oxidative stress inhibit the proliferation of normal ARPE-19 cells, increase the expression of VEGF-A and reduce the expression of Akt.