[A monoclonal antibody-based icELISA for puerarin].

Puerarin was conjugated with bovine serum albumin(BSA) and ovalbumin(OVA) by periodate oxidation to serve as the immunogen and coating antigen, respectively. BALB/c mice were immunized with puerarin-BSA according to the routine immunization procedure, and the titer and specificity of serum were detected after three immunization. After booster immunization, mouse spleen lymphocytes were fused with mouse myeloma cells, and 24 hybridoma cell lines of the monoclonal antibodies against puerarin were screened by monoclonal antibody screening technique. Ascites was prepared and purified. The cross-reactivity of monoclonal antibody(mAb) M1 with 4'-methoxy puerarin, daidzin, puerarin-6″-O-xyloside, daidzein, mirificin, 3'-methoxy puerarin, and 3'-hydroxy puerarin was 239.84%, 112.18%, 67.89%, 58.28%, 22.37%, 0.40%, and 0.20%, respectively, and those with other analogs such as baicalein and baicalin were all less than 0.10%. The IC_(50) and the working range of the indirect competitive enzyme-linked immunosorbent assay(icELISA) for puerarin were 44.80 ng·mL~(-1) and 8.20-292.30 ng·mL~(-1), respectively. The average recovery was 91.95%-98.20% with an RSD in the range of 0.70%-2.60%. The content of puerarin in different Puerariae Lobatae Radix samples was determined with icELISA and validated by UPLC-MS. The correlation between data obtained from icELISA and UPLC-MS was 0.999 0, indicating that icELISA is suitable for the rapid detection of puerarin in Puerariae Lobatae Radix samples.

Effect of the regimen of Gaoshan Hongjingtian on the mechanism of poly (ADP-ribose) polymerase regulation of nuclear factor kappa B in the experimental diabetic retinopathy.

BACKGROUND: Poly(ADP-ribose) polymerase (PARP) plays an important role in the death of retinal capillary cells in diabetic retinopathy (DR) partly via its regulation of nuclear factor kappa B (NF-κB). The current study investigated the effect of the regimen of Gaoshan Hongjingtian (RG) on the mechanism of PARP regulation of NF-κB, and demonstrated the possible impact of the RG and Gaoshan Hongjingtian (Rhodiola sachalinensis, RS) on diabetic retinopathy. METHODS: Wistar rats were made diabetic by administering streptozotocin. They were then assigned to three groups at random. After 2 months, the three groups of these diabetic rats were treated with RS or RG, or untreated. Analyses of expression levels of PARP, NF-κB, and intercellular adhesion molecule-1 (ICAM-1) in the retinas of rats in different groups were performed by Western blotting and immunohistochemical assays, and mRNA levels of NF-κB and ICAM-1 were determined by real-time polymerase chain reaction (PCR). In addition, the basement membranes of capillaries in the rats' retinas were observed using electron microscopy, and diabetes-induced capillary degeneration (ghost pericytes and acellular capillaries) were quantitated. RESULTS: From the third month after the injection of streptozotocin, the diabetic rats were given daily RG, RS or tap water separately. The diabetic rats failed to gain weight compared with normal age-matched rats, whereas their glycated hemoglobin levels were significantly increased. After 5 months, the mRNA levels of NF-κB and ICAM-1 and the protein expression of PARP, NF-κB, and ICAM-1 were significantly increased in the retinas of diabetic rats in the untreated group compared with the nondiabetic controls. After 8 months, the number of degenerated retinal capillaries (ghost pericytes and acellular capillaries) was significantly increased in the diabetic rats in the untreated group compared with normal age-matched rats. RG and RS inhibited diabetes-induced over-expression of PARP, NF-κB, and ICAM-1 in the retinas of diabetic rats at the end of 5-month diabetic duration. Treatment using RG and RS significantly inhibited increases in the number of acellular capillaries and pericyte ghosts and suppressed the basement membrane thickening in the retinas of rats with diabetes for 8 months compared with the control diabetic rats. CONCLUSIONS: These results indicate that PARP plays an important role in the pathogenesis of diabetic retinopathy. RS and RG may have acted on the mechanism of PARP regulation of NF-κB, which suppressed the expression of NF-κB and ICAM-1, and led to the inhibition of retinal capillary degeneration.

[Transpupillary thermotherapy for circumscribed choroidal hemangiomas].

OBJECTIVE: To study the effects of transpupillary thermotherapy (TTT) on circumscribed choroidal hemangiomas (CCH). METHODS: It was a retrospective cases series study. One hundred and fourteen cases (114 eyes) in out-patient department were enrolled in the study with 80 male cases and 34 female cases. The age ranged from 16 to 71 yrs with an average at 44 years. Single eye was affected in all cases. Indirect ophthalmoscope, fundus fluorescein angiography (FFA), ultrasound (B) were used for the examination. Treatment was delivered via slit lamp by using an 810 nm diode laser (Irises) with spots or composed spots (larger tumor) covering the whole tumor. If the tumor was located or partly located in the fovea, laser spot should be avoided in the fovea and papillo-macular bundle. The laser power should be lesser (light gray color) in parafovea area. Routine laser spot (gray-white color) was used in perifovea area. A width of 200 µm out of disk border should not be placed by laser spot in paradisk tumor cases. RESULTS: The visual acuity (VA) ≥ 0.5 of pre-and post-treatment was 17 eyes (14.9%) and 27 eyes (23.7%), respectively. Pre-treatment, the retinal detachment extended in the inferior part in 27 cases (23.7%) except on the tumor surface. The tumor was located at macular area in 67 cases (58.8%), around the disc in 35 cases (30.7%), at the temporal arcuate in 12 cases (10.5%). In our cases, VA maintenance with fluid absorption was noted in 76 cases (66.7%) after the treatment. VA improvement with fluid absorption in 28 case (24.6%). Both VA maintenance and improvement (rate of efficiency) with fluid absorption were 91.2%. Fluid non-absorption or VA decline was regarded as unsuccessful in 10 cases (8.8%). The average thickness of 32 cases tumor pre- and post-treatment was (3.90 ± 1.15) mm and (2.41 ± 1.30) mm. Twenty seven cases were followed up for ≥ 1 year (mean 22.8 months) with rate of efficiency at 81.5%. The complications (9 eyes, 7.9%) contained minor macular hemorrhage in 2 eyes, macular pucker in 3 eyes, macular edema and cystoid macular edema in 2 eyes, retinal branch occlusion in 1 eye, and arcuate scotoma in 1 eye. CONCLUSIONS: The treatment of TTT on CCH, whether the tumor located around-disc or in foveal area with exudative fluid could get successful in the majority of cases. TTT is one of the treatments worth doing owing to fewer complications, less expenses and easy-doing with definite and persistent effect.

Treatment of retinal vein occlusion in rabbits with traditional Chinese medicine Fufang XueShuan Tong.

BACKGROUND: Retinal vein occlusion (RVO) is one of the most common causes of visual loss. Many approaches have been tried to treat central retinal vein occlusion (CRVO), and branch retinal vein occlusion (BRVO) with various results. However, there is no defined protocol and limited evidence to support the interventions currently used. The aim of this study was to assess the efficacy of the traditional Chinese medicine Fufang XueShuan Tong (FXST) in treating experimentally created RVO. METHODS: RVO model was first induced in forty-four pigmented rabbits through photocoagulation following injection of rose Bengal. The rabbits were divided into four groups based on the dose of FXST administered (212 mg/kg, 424 mg/kg, 848 mg/kg and control group). The rabbits were observed for four weeks after the procedure, using color fundus photography, fundus fluorescein angiography and electroretinogram examination. Vascular endothelial growth factor (VEGF), interleukin-6 and nitric oxide (NO) levels in the vitreous and histopathologic evaluation were monitored. RESULTS: The obstructed vessels in the treatment groups reopened or anastomosed faster than those in the control group (P < 0.05). The amplitude of maximum b wave and the oscillatory potential were significantly higher in the treatment groups than in the control group (P < 0.01). At both two weeks and four weeks, VEGF and IL-6 levels in the vitreous were significantly decreased in the treatment groups (P < 0.01), while NO levels were significantly elevated (P < 0.01). At the same time, histopathologic evaluation showed different retinal neuroepithelium structures in the different groups. Immunoreactivity of VEGF was greater in the control group than in the treatment groups. CONCLUSION: FXST was helpful in reconstructing retinal vessels in the RVO model, protecting retinal structures and improving visual function, and could inhibit the neovascular factor.

[Clinical features and treatment of choroidal metastasis].

OBJECTIVE: To assess the clinical features and management of choroidal metastasis. METHODS: Fundus examination was performed in 49 patients (66 eyes) with choroidal metastasis. Fundus fluorescein angiography (FFA) was performed in 44 cases, combined with indocyanine green angiography (ICGA) examination in 12 cases. B-scan ultrasound examination was performed in 8 cases. Transpupillary thermotherapy (TTT) was performed in 24 eyes, combined with photo-dynamic therapy in one eye. Plaque radio-therapy was used in one eye. The parameters of treatment for TTT were 1.2 - 3 mm spot size, 450 - 1000 mV, 60 s; 2 sessions of TTT in 2 eyes and 3 sessions in 3 eyes. RESULTS: Fourteen cases were male and 35 cases were female. Both eyes were affected in 17 cases (34.7%). Age ranged from 23 - 74 years old with an average of 47 years. The visual acuity was 0.05 or less in 13 eyes; 0.06 - 0.2 in 22 eyes and 0.3 or more in 31 eyes. Primary tumours were found in 40 cases (81.6%) (surgical excision in 25 cases), consisting of breast carcinoma in 16 cases (32.7%), lung carcinoma in 14 cases (28.6%), hepatoma and cholangiocarcinoma in 3 cases, colon and stomach carcinomas in 3 cases, gynecologic appendix carcinoma (including 1 case of ovarian mucous cyst adenocarcinoma) in 2 cases, nasopharyngeal adenocarcinoma in 1 case, vertebra tumor in 1 case, undetected in 5 cases (10.2%) and under detection in 4 cases (8.2%). The fundus had 1 lesion in 58 eyes (58/66 = 87.8%), 2 lesions in 4 eyes (4/66 = 6.0%), 3 or more lesions in 2 eyes (including 7 lesions in 1 eye). According to the location and development status of the lesions, they could be divided into solitary type, 39 eyes (39/66 = 59.1%); diffuse type, 19 eyes (19/66 = 28.8%); and early type, 8 eyes (8/66 = 12.1%). FFA examination: early stage lesions showed hypofluorescence and later stage lesions showed moderate to strong hyperfluorescence. In 8 cases of solitary lesions, the size of the lesion measured by B-scan averaged 11.5 mm x 10.5 mm x 3.6 mm with the maximal height at 4.9 mm. The tumor became flattened and vision remained stable at 3 months after plaque radiotherapy in 1 case. Three cases were followed-up for 2, 3, and 4 months after TTT treatment. The lesions remained stable with vision unchanged or slightly decreased. CONCLUSIONS: The choroidal metastasis has specific clinical features. The classification of metastatic lesions into solitary, diffuse and early types is helpful for the evaluation of the disease process. The primary tumor can be found in 80% of cases. The most common primary cancer is breast carcinoma, followed by lung carcinoma. These two cancers account for 75% of primary tumors. In solitary type and early type lesions, TTT combines with systemic treatment could result in regression of lesions, saving of vision and improvement of the life quality.

[The effect of retrobulbar injection with anisodamine on ocular blood velocity in anterior ischemic optic neuropathy].

OBJECTIVE: To study the effect of retrobulbar injection with anisodamine on ocular blood velocity in anterior ischemic optic neuropathy (AION). METHODS: 15 minutes before injection, 15 minutes and 1 hour after injection, the blood flow velocity in 39 cases (39 eyes) diagnosed as AION was measured and analyzed by CDI, timed average maximum velocity, peak systolic velocity, end diastolic velocity, resistance index and pulsatility index of OA, CRA and PCA were recorded. RESULTS: Compared with the normal eyes, each parameter of SPCA was statistically significant. 15 minutes and 1 hour after post ocular injection of anisodamine, every parameter examined in the experiment was increased except resistance index compared with pre-injection (P < 0.01). CONCLUSIONS: retrobulbar injection with anisodamine can effectively improve flow velocity of nasal and tempo short posterior ciliary artery and central retinal artery.