Objective To establish a human luminal breast cancer stem cell(BCSC)model and investigate the inhibitory effects of astragaloside Ⅳ(AS-Ⅳ)on BCSC growth.Methods MCF-7 breast cancer cells were cultured in stem cell-specific medium to induce BCSC formation.The BCSCs were then divided into a blank control group and an AS-Ⅳ treatment group,both groups were given PBS or AS-Ⅳ treatment.Morphological changes were observed after intervention.The therapeutic efficacy of AS-Ⅳ was evaluated using 3D spheroid formation and cell viability assays.Transcriptomic profiling and gene expression analysis were performed to elucidate the underlying mechanisms.Results Compared with the MCF7 breast cancer cells,MCF7 breast cancer stem cell mammospheres exhibited accelerated growth(P＜0.01)and significantly increased expression of the stemness marker ALDH1A1(P＜0.01).Further comparison with the blank control group revealed that astragaloside Ⅳ(AS-Ⅳ)treatment significantly inhibited MCF7 breast cancer stem cell proliferation(P＜0.001)and slowed mammosphere growth(P＜0.01).Transcriptomic analysis demonstrated that differentially expressed genes(DEGs)induced by stem cell modeling and AS-Ⅳ intervention were enriched in the cellular senescence signaling pathway.AS-Ⅳ intervention substantially increased the number of SA-β-gal-positive cells(P＜0.01).RT-PCR analysis confirmed that AS-Ⅳsignificantly upregulated mRNA expression of IL-1α(P＜0.01),P21(P＜0.001),and P53(P＜0.05)in MCF7 breast cancer stem cells.Conclusion Astragaloside Ⅳ suppresses the growth of human luminal breast cancer stem cells by inducing cellular senescence.

Objective:To evaluate the safety and efficacy of 27G micro-incision vitrectomy surgery (MIVS) combined with intravitreal injection of ranibizumab (IVR) in the treatment of retinopathy of prematurity (ROP) with early intervention failure.Methods:Retrospective case series was performed. Fourteen eyes (11 infants) with ROP who underwent 27G MIVS combined with IVR were included from March 2016 to January 2018 in Shenzhen Eye Hospital. Among them, there were 5 males with 7 eyes, 6 females with 7 eyes. The average gestational age of the infants was 28.12±0.90 weeks; the average birth weight was 1 023.64±200.96 g. Before the early clinical intervention, 1 infant (2 eyes) had ROP in zone Ⅰ stage 3 with plus disease, 8 infants (10 eyes) had ROP in zone Ⅱ stage 3 with plus disease, and 2 infants had ROP in aggressive posterior ROP. Six eyes underwent laser photocoagulation, while 8 eyes received laser therapy combined with IVR. Six eyes of stage 4A ROP and 8 eyes in stage 4B. Retinal detachment was detected with a mean of 10.44±9.21 weeks. At the time of surgery, the average post-conceptional age was 48.02±8.09 weeks. All the affected eyes were treated with standard sclera with three incisions 27G MIVS. During the operation, only local vitrectomy was performed to release and clear fibroascular proliferation in the optic disc, anterior macular area and pericristal area. After surgery, 10 mg/ml of ranibizumab 0.03 ml was injected into the vitreous cavity. The average follow-up time was 23.36±8.34 months. The primary objectives were the condition of retinal reset, ROP progression control and complications.Results:All patients had uneventful surgeries with an average duration of 32.86±9.35 mins. Of the 14 eyes, 12 eyes (85.71%) were controlled, 8 eyes (57.14%) had a good rearrangement of macular structure, while 4 eyes with macular traction. Two eyes had ROP progression, recurrence of retinal detachment, posterior synechia. Complicated cataract was in 1 eye. Proliferative vitreoretinopathy and retinal detachment was in 1 eye after 7 months the operation.Conclusion:27G MIVS combined with IVR is a safe and effective treatment for ROP with early clinical intervention failure.

Objective:To explore the feasibility to establish a novel retinal ischemia reperfusion (RIR) injury model by applying pars plana vitrectomy (PPV) combined with retinal vascular counterpulsation in the central retinal artery occlusion (CRAO) model of New Zealand rabbit.Methods:Twenty New Zealand adult rabbits were randomly divided into two groups by random number table, 10 for each group.Rabbits in the laser group were treated with retinal laser photocoagulation only, while rabbits in the counterpulsation group were treated with PPV combined counterpulsation.The right eye of each New Zealand rabbit was used as the experimental eye and the left eye was used as the normal control (the normal control group). Fundus fluorescence angiography (FFA) , oxygen partial pressure (PO 2) in vitreous cavity was performed to assess the recovery status of perfusion.Scotopic 3.0 oscillatory potentials (OPs) in electroretinogram (ERG) were used to evaluate the retinal function, and retinal pathological sections were used to evaluate the structural changes in the retina.The use and care of the animals complied with the Statement of the Association for Research in Vision and Ophthalmology (ARVO), and the study was approved by the Animal Research Committee of Zhongshan Ophthalmic Center, Sun Yat-sen University. Results:In the counterpulsation group, retinal reperfusion was observed during counterpulsation processure.FFA examination at 2 hours after counterpulsation showed reperfusion of retinal blood stream in all the eyes.Retinal artery filling, followed by venous filling was observed in the early stage, with no delay in filling and no interruption of blood flow.The percentage of vitreous PO 2 was significantly different among the counterpulsation group, the laser group and the normal control group at different time points ( Fgroup=330.87, P<0.001; Ftime=985.70, P<0.001). The percentages of vitreous PO 2 in the counterpulsation group at different time points was (18.67±6.29)%, (38.82±1.48)%, (57.33±4.25)%, (84.51±3.91)% and (89.20±2.97)%, which were significantly higher than that in the laser group ([23.24±1.95]%, [31.44±3.29]%, [40.21±3.05]%, [43.65±3.82]% and [58.07±2.93]%) (all at P<0.05). The percentage of OPs was significantly different among the counterpulsation group, the laser group and the normal control group at different time points ( Fgroup=164.09, P<0.001; Ftime=447.91, P<0.001). The percentages of OPs of counterpulsation group at different time points after laser and after PPV were (47.23±2.73)%, (70.79±3.09)%, (78.39±3.63)%, (76.69±4.08)% and (82.18±1.78)%, which were significanthy higher than that in the laser group ([46.83±2.89]%, [55.32±1.58]%, [51.08±4.02]%, [52.32±6.59]% and [53.46±6.46]%) ( all at P<0.05). There was a lesser damage in inner retinal structure in pathological section in the counterpulsation group.The myelinated nerve fiber layer (MFL) was loose and a mass of vacuolar changes were observed in MFL.The structure of MFL, inner plexiform layer, inner and outer plexiform layer in the laser group were disordered, and the Müller cell nerve fibers were destroyed in the laser group. Conclusions:A novel type of RIR injury model can be established by applying PPV combined with counterpulsation in the CRAO model of New Zealand rabbit.

Objective To explore the reaction pattern rules of mouse retinal ganglion cells potential under different wavelengths of light stimulation.Methods Thirty SPF grade 3-week-old C57BL/6 mice were used for ex vivo whole mount retina preparation.The cells firing activities were recorded on patch clamp system with on cell touch mode under stimulation of 400 nm,580 nm and white light,respectively.According to different reactions to different light stimulation,the cells were classified into 400 nm sensitive RGC,580 nm sensitive RGC and color vision insensitive RGC.Then the cells were further classified according to light ON type,light ON/OFF type or light OFF type.The RGC's baseline firing pattern (baseline firing frequency,burst firing frequency) and light activation firing pattern (response pattern,light response firing frequency,light response firing amplification) were compared among different RGC classifications.Results Eighty-two RGCs were recorded in total.The frequency of spontaneous firing activity ranged from 0.00 Hz to 32.33 Hz among different RGCs.400 nm sensitive RGCs were 52 (63.41％),580 nm sensitive RGCs were 29(35.37％) and color vision insensitive RGC was 1 (1.22％).OFF type RGC was the main cell type in 400 nm sensitive group (36.29％),and ON/OFF type RGC was the main cell type in 580 nm sensitive group (34.48％).The firing amplification in 580 nm sensitive RGC was (22.93±10.23) Hz,which was significantly higher than (14.44± 10.11) Hz in 400 nm sensitive RGC (t =4.060,P =0.044).The firing amplification in 580 nm sensitive ON type RGC was (24.17±8.98)Hz,which was significantly higher than (11.12±10.35)Hz in 400 nm sensitive ON type RGC (t =5.373,P =0.021).Conclusions There is no specific firing pattern rules among different light sensitive RGCs.In the future,artificial color vision may be achieved through personalized electric stimulation and learning feedback strategy.

Objective To explore the reaction pattern rules of mouse retinal ganglion cells potential under different wavelengths of light stimulation. Methods Thirty SPF grade 3.week.old C57BL/6 mice were used for ex vivo whole mount retina preparation. The cells firing activities were recorded on patch clamp system with on cell touch mode under stimulation of 400 nm,580 nm and white light,respectively. According to different reactions to different light stimulation, the cells were classified into 400 nm sensitive RGC, 580 nm sensitive RGC and color vision insensitive RGC. Then the cells were further classified according to light ON type,light ON/OFF type or light OFF type. The RGC's baseline firing pattern ( baseline firing frequency,burst firing frequency) and light activation firing pattern (response pattern,light response firing frequency,light response firing amplification) were compared among different RGC classifications. Results Eighty.two RGCs were recorded in total. The frequency of spontaneous firing activity ranged from 0. 00 Hz to 32. 33 Hz among different RGCs. 400 nm sensitive RGCs were 52(63. 41％),580 nm sensitive RGCs were 29(35. 37％) and color vision insensitive RGC was 1(1. 22％). OFF type RGC was the main cell type in 400 nm sensitive group (36. 29％),and ON/OFF type RGC was the main cell type in 580 nm sensitive group (34. 48％). The firing amplification in 580 nm sensitive RGC was (22. 93±10. 23)Hz,which was significantly higher than (14. 44±10. 11)Hz in 400 nm sensitive RGC (t=4. 060,P=0. 044). The firing amplification in 580 nm sensitive ON type RGC was (24. 17±8. 98)Hz,which was significantly higher than (11. 12±10. 35)Hz in 400 nm sensitive ON type RGC (t=5. 373,P=0. 021). Conclusions There is no specific firing pattern rules among different light sensitive RGCs. In the future, artificial color vision may be achieved through personalized electric stimulation and learning feedback strategy.