Betaine addition to the diet alleviates intestinal injury in growing rabbits during the summer heat through the AAT/mTOR pathway

Background The aim of this experiment was to investigate the effect of different levels of betaine(Bet)inclusion in the diet on the intestinal health of growing rabbits under summer heat.A total of 100 weaned Qixing meat rabbits aged 35 d with body weight of 748.61±38.59 g were randomly divided into 5 treatment groups:control group(basal diet)and Bet groups(basal diet+0.75,1.0,1.5 or 2.0 g/kg Bet).The average daily temperature in the rabbitry during the experiment was 30.48°C and the relative humidity was 69.44%.Results Dietary addition of Bet had no significant effect on growth performance and health status of growing rabbits(P>0.05),but it increased ileal secretory immunoglobulin A content compared to the control under summer heat(P<0.05).Addition of 0.75 g/kg Bet up-regulated jejunal IL-4,down-regulated ileal TNF-αexpression(P<0.05).The addition of 1.0 g/kg Bet increased the villi height(VH)in the jejunum(P<0.05).Serum glucose levels were reduced,and the expression of SLC6A20 was up-regulated in jejunum and ileum of rabbits fed with 1.5 g/kg Bet(P<0.05).When added at 2.0 g/kg,Bet reduced serum HSP70 content,increased jejunal VH,and up-regulated duodenal SLC7A6,SLC38A2,mTOR and 4EBP-2 expression(P<0.05).Correlation analysis revealed that intestinal mTOR expression was significantly and positively correlated with SLC7A6,SLC38A2,SLC36A1 and IL-4 expression(P<0.05).Conclusions Dietary addition of Bet can up-regulate the expression of anti-inflammatory factors through the AAT/mTOR pathway,improve the intestinal immune function,alleviate intestinal damage in growing rabbits caused by summer heat,and improve intestinal health.

Genome-edited rabbits:Unleashing the potential of a promising experimental animal model across diverse diseases

Animal models are extensively used in all aspects of biomedical research,with substantial contributions to our understanding of diseases,the development of pharmaceuticals,and the exploration of gene functions.The field of genome modification in rabbits has progressed slowly.However,recent advancements,particularly in CRISPR/Cas9-related technologies,have catalyzed the successful development of various genome-edited rabbit models to mimic diverse diseases,including cardiovascular disorders,immunodeficiencies,agingrelated ailments,neurological diseases,and ophthalmic pathologies.These models hold great promise in advancing biomedical research due to their closer physiological and biochemical resemblance to humans compared to mice.This review aims to summarize the novel gene-editing approaches currently available for rabbits and present the applications and prospects of such models in biomedicine,underscoring their impact and future potential in translational medicine.

Effects of Incorporating Mango (Mangifera indica), Avocado (Persea americana) and Guava (Psidium guajava) Leaves Powder in the Ration on Growth Characteristics in Rabbits

A study conducted on burgundy rabbits evaluated the effects of rations containing mango (Mangifera indica), avocado (Persea americana) and guava (Psidium guajava) powder. For this purpose, 36 rabbits (20 males and 16 females) aged 60 to 70 days with an average weight of 1012 ± 133 g were divided randomly into 4 equal groups (T0, T1, T2 and T3) of 9 animals each according to body weight (bw). The rabbits were divided throughout the trial, the animals were fed, those of group T0 (control) with a standard feed without any type of leaf powder, while those of groups 1, 2 and 3 received the standard feed supplemented with Mangifera indica, Persea americana and Psidium guajava leaves powder at the rate of 0.5% respectively. After two months of treatment, the animals were fasted for 12 hours (20 h - 8 h) and sacrificed to evaluate carcass and digestive organ weights. The results revealed that the values of feed intake, live weight and weight gain were non-significantly (p > 0.05) increased in all animals regardless of the ration in reference to the control. The different leaves powder induced an increase in carcass yield in rabbits but this increase was only significant (p M. indica leaf powder (51.03 ± 0.24) as compared to the control (48.63 ± 0.74). Gut length was significantly increased in animals fed rations containing Psidium guajava and Persea americana leaves powder compared to the control. A significant increase in gut weight was observed in animals fed the Persea americana leaf supplemented ration. In general, mango leaf powder performed better than other types of powders.

An Improved Artificial Rabbits Optimization Algorithm with Chaotic Local Search and Opposition-Based Learning for Engineering Problems and Its Applications in Breast Cancer Problem

Artificial rabbits optimization(ARO)is a recently proposed biology-based optimization algorithm inspired by the detour foraging and random hiding behavior of rabbits in nature.However,for solving optimization problems,the ARO algorithm shows slow convergence speed and can fall into local minima.To overcome these drawbacks,this paper proposes chaotic opposition-based learning ARO(COARO),an improved version of the ARO algorithm that incorporates opposition-based learning(OBL)and chaotic local search(CLS)techniques.By adding OBL to ARO,the convergence speed of the algorithm increases and it explores the search space better.Chaotic maps in CLS provide rapid convergence by scanning the search space efficiently,since their ergodicity and non-repetitive properties.The proposed COARO algorithm has been tested using thirty-three distinct benchmark functions.The outcomes have been compared with the most recent optimization algorithms.Additionally,the COARO algorithm’s problem-solving capabilities have been evaluated using six different engineering design problems and compared with various other algorithms.This study also introduces a binary variant of the continuous COARO algorithm,named BCOARO.The performance of BCOARO was evaluated on the breast cancer dataset.The effectiveness of BCOARO has been compared with different feature selection algorithms.The proposed BCOARO outperforms alternative algorithms,according to the findings obtained for real applications in terms of accuracy performance,and fitness value.Extensive experiments show that the COARO and BCOARO algorithms achieve promising results compared to other metaheuristic algorithms.

Radiosensitivity of β-elemene on rabbit VX2 renal transplant carcinoma model

Objective To investigate the effects of low dosage of β-elemene on the radiosensitivity of rabbit VX2 renal transplant carcinoma model. Methods We took the rabbit VX2 renal transplant carcinoma as the model. Experimental rabbits were divided into three groups: the control group, the radiation group, and the radiation +β-elemene (radiosensitivity) group. The change of tumor was observed by Spiral CT and B ultrasound to compare its regrowth period. The tumor was measured by light microscopy and electron microscopy. Results The tumor in radiosensitivity group was restrained obviously and the sensitization enhancement ratio (SER) of β-elemene was 1.89. Different apoptosis was observed under transmission electron microscopy. Conclusion Low dosage β-elemene can enhance the radiosensitivity of rabbit VX2 renal transplant carcinoma model and induce the apoptosis of tumor cells, but the mechanism needs further study. It promotes apoptosis in mechanisms in vitro.

Intravitreal slow-release dexamethasone alleviates traumatic proliferative vitreoretinopathy by inhibiting persistent inflammation and Müller cell gliosis in rabbits

AIM:To evaluate the effects of intravitreal slow-release dexamethasone on traumatic proliferative vitreoretinopathy(PVR)and Müller cell gliosis and preliminarily explored the possible inflammatory mechanism in a rabbit model induced by penetrating ocular trauma.METHODS:Traumatic PVR was induced in the right eyes of pigmented rabbits by performing an 8-mm circumferential scleral incision placed 2.5 mm behind the limbus,followed by treatment with a slow-release dexamethasone implant(Ozurdex)or sham injection.Left eyes were used as normal controls.The intraocular pressure(IOP)was monitored using an iCare tonometer.PVR severity was evaluated via anatomical and histopathological examinations every week for 6wk;specific inflammatory cytokine and proliferative marker levels were measured by quantitative real-time polymerase chain reaction,Western blot,protein chip analysis,or immunofluorescence staining.RESULTS:During the observation period,PVR severity gradually increased.Intense Müller cell gliosis was observed in the peripheral retina near the wound and in the whole retina of PVR group.Ozurdex significantly alleviated PVR development and Müller cell gliosis.Post-traumatic inflammation fluctuated and was persistent.The interleukin-1β(IL-1β)mRNA level was significantly upregulated,peaking on day 3 and increasing again on day 21 after injury.The expression of nod-like receptor family pyrin domain containing 3(NLRP3)showed a similar trend that began earlier than that of IL-1βexpression.Ozurdex suppressed the expression of IL-1β,NLRP3,and phosphorylated nuclear factor-kappa B(NF-κB).The average IOP after treatment was within normal limits.CONCLUSION:The present study demonstrates chronic and fluctuating inflammation in a traumatic PVR rabbit model over 6wk.Ozurdex treatment significantly inhibites inflammatory cytokines expression and Müller cell gliosis,and thus alleviates PVR severity.This study highlights the important role of IL-1β,and Ozurdex inhibites inflammation presumably via the NF-κB/NLRP3/IL-1βinflammatory axis.In summary,Ozurdex provides a potential therapeutic option for traumatic PVR.

Effect of taxifolin on cold-shock damages in spermatozoa in rabbits

Objective:To investigate the effect of taxifolin added to rabbit semen on freezing-induced cold-shock damages in spermatozoa.Methods:Semen was collected from six adult New Zealand rabbits once a week by artificial vagina.The collected semen was pooled at 38℃ and divided into four equal volumes.They were diluted with 0,50,100 and 200μM taxifolin-containing Tris+egg yolk extender at 38℃ and their temperatures were lowered to 4℃.Following equilibration,semen drawn into 0.25 mL straws were frozen in an automatic semen freezing device and stored in liquid nitrogen container at-196℃.Samples were thawed in 38℃ water for 25 s and the analyses of motility,kinematic parameters,morphological deformities,changes in membrane integrity,mitochondrial membrane potential,dead-live ratio,acrosomal damages and as well as oxidative stress analyses were performed in semen.Results:Addition of 50μM taxifolin significantly improved motility(total,progressive,rapid and static),high mitochondrial membrane potential and the ratios of spermatozoa with acrosomal damage compared to the control group.Compared to the control group,malondialdehyde(MDA)levels in the 50 and 100μM taxifolin groups were significantly lower,while the MDA level was high and viable spermatozoa ratio was low in the 200μM taxifolin group.There were no significant differences between the groups in terms of kinematic parameters,morphological deformities,membrane integrity and antioxidant levels.Conclusions:The low dose of taxifolin(50μM)has a positive effect and the high dose(200μM)has a negative effect.Therefore,it is concluded that the addition of low-dose(50μM)taxifolin to the extenders would be a useful additive in reducing cold-shock damage that occurs during freezing of rabbit semen.

A novel approach for 25-gauge transconjunctival sutureless vitrectomy to evaluate vitreous substitutes in rabbits

AIM:To improve the standard three-port vitrectomy for establishing and evaluating an endotamponade model in rabbits.METHODS:Three ports were prepared near the third eyelid of rabbits,and the infusion port was placed at the inferior nasal quadrant with the inserted cannula linking with a self-designed handheld rigid infusion catheter.All right eyes of rabbits underwent a modified 25-gauge vitrectomy and were subsequently filled with balanced salt solution,silicone oil,and eight-arm polyethylene glycols(8-arm PEGs)hydrogel separately for comparison.Ophthalmic examinations were performed regularly to record the changes after the surgery.RESULTS:Successful vitrectomy was achieved among 44 chinchilla rabbits.The mean operation time was 4.51±1.25min.Four eyes(9.1%)presented limited lens touch and two eyes(4.5%)showed retinal touch during surgery.Incision leakage was found in three eyes(6.8%)after surgery.There was no endophthalmitis,hemorrhage,or retinal detachment during the observation period and ophthalmic examinations after the implantation of vitreous substitutes.CONCLUSION:The modified technique of the standard vitrectomy applied in the endotamponade model in rabbits shows excellent safety and practicality.

Analysis and Identification of miRNA Expression in the Skeletal Muscle of Sichuan White Rabbits

[Objectives]miRNAs play an important role in the proliferation and differentiation of different myoblasts.This study was conducted to elucidate the complex genetic mechanisms that affect the meat production performance of Sichuan white rabbits and reveal the regulatory role of miRNAs in their muscle growth and meat quality formation.[Methods]Three constructed skeletal muscle libraries of Sichuan white rabbits aged six months were sequenced by the solexa technology to identify known miRNAs,predict new miRNAs and construct an expression profile of muscle miRNAs.[Results]A total of 511 known miRNAs and 42 miRNAs were detected in 34089472 pure sequences,and the proportion of miRNAs with a length of 22 nt was the highest.The number of known miRNA sequences accounted for 71.38%of pure sequences,which was much higher than the proportion of other types of RNAs.The proportion of sequences from exons was 0.38%,indicating a low degree of mRNA degradation in the samples.Base U had the highest proportion at the first position,and the bases with the highest proportions at positions 8 and 10 were U and A,respectively.Muscle-specific miRNAs(miR-1,miR-133,and miR-206)ranked in the top 10 in terms of expression level.The number and expression levels of new miRNAs were lower than those of known miRNAs.The length distribution,base bias at different positions and expression profile characteristics of miRNAs might be related to the biological function of miRNAs in regulating muscle proliferation and differentiation and the action mechanisms with target genes.[Conclusions]The identification and expression of miRNAs in muscle tissues of Sichuan white rabbits will help to understand the complex molecular mechanisms of meat production performance and provide a theoretical basis for the functional research of miRNAs in meat rabbits.

β-Elemene reduces the progression of atherosclerosis in rabbits

The present study aimed at investigating the possible effects offl-elemene on the progression of atherosclerosis in a rabbit model. The rabbit atherosclerosis model was established by the combination of balloon angioplasty-induced endothelial injury and an atherogenic diet fed to the rabbits. New Zealand White rabbits were randomly divided into four groups （8/group）： the normal control group （fed with normal chow diet）, and three experimental groups, placebo group, atorvastatin group, and β-elemene group （received the atherogenic diet）. After two weeks on the diet, the three experimental groups underwent balloon injury at right common carotid artery and were treated with drugs or placebo for five weeks. Serum lipids were measured, Carotid artery lesions were isolated for histological and immunohistochemical analysis. In vitro, RAW264.7 macrophages were pretreated with β-elemene and ox-LDL for 24 h and the viability of macrophages was assayed using the MTT method. TNF-a and IL-6 were also determined. Compared with the control group, the thickness of the atherosclerosis lesion in the placebo group was significantly increased; The thickness the drug treatment groups were significantly decreased, compared with that of the placebo group. The infiltration of macrophage was markedly reduced in the β-elemene group compared with that of the placebo group,β-Elemene treatment also reduced the levels of TC, TC, and LDL-C, compared with the placebo group, β-elemene decreased the TNF-a and IL-6 levels in vitro. In conclusion, our results demonstrated that β-elemene retarded the progression of atherosclerosis in vivo and in vitro, which may be related to the capacity of β-elemene to reduce the infiltration of macrophages and suppress inflammatory factors.

Experimental Study on the Repair Effect of Xianlinggubao Capsule on Osteoporotic Vertebral Compression Fracture in Rabbits

[Objectives]To observe the effect of Xianlinggubao Capsule on osteoporotic vertebral compression fracture(OVCF)in rabbits and the influence mechanism of the repair of fractures.[Methods]Female June age 30 rabbits were randomly divided into control group,model control group and Xianlinggubao group.After bilateral ovariectomy,the model control group and Xianlinggubao group were injected with dexamethasone continuously for 4 weeks,and then the OVCF compound model was established by surgery.The Xianlinggubao group was treated with Xianlinggubao at a dose of 300 mg/(kg·d)for 60 d,while the blank control group and the model control group were treated with the same amount of normal saline for 60 d.The number of blood vessels and the expression of bone morphogenetic protein-2(BMP-2)were detected by immunohistochemical staining and the bone mineral density(BMD)in the callus of the third lumbar fracture area of rabbits was measured.The content of serum phosphorus(P),alkaline phosphatase(ALP)and total calcium(TCa)in rabbit venous blood were measured by automatic biochemical analyzer.The content of vascular endothelial growth factor(VEGF)and platelet-derived growth factor(PDGF)in rabbit venous blood were measured by ELISA kit.[Results]The number of blood vessels and the expression of BMP-2 in the callus of the third lumbar fracture area of rabbits was high in Xianlinggubao group,the content of serum P,ALP,TCa,VEGF and PDGF was obviously increased,BMD was obviously increased,the bone microstructure of the third lumbar vertebrae fracture area of rabbits was basically restored.Compared with the model control group(P<0.05),the difference was statistically significant.[Conclusions]Xianlinggubao Capsule can increase calcium and phosphorus deposition,promote the formation of blood vessels in the fracture area of OVCF in rabbits,and have a strong repair effect on OVCF in rabbits.

β-elemene通过调节耐药特异性miRNA逆转乳腺癌细胞耐药性的机制研究

目的:探讨β-榄香烯逆转乳腺癌细胞耐药性的机制,明确其抗肿瘤作用。方法:通过向MCF-7细胞株培养基内逐渐增加临床上最常用于乳腺癌化学治疗的药物多西紫杉醇(Doc)或阿霉素(Adr),逐渐提高培养基中药物浓度的方法成功地构建出亲本细胞株MCF-7中耐药亚型,选择乳腺癌阿霉素耐药细胞(MCF-7/Adr)和乳腺癌多西紫杉醇(MCF-7/Doc)耐药细胞株,采用MTT-cytotoxic、miRNA微阵列、实时定量PCR、双荧光素酶活性测定等方法,研究β-elemene对乳腺癌耐药细胞生存率、耐药特异性miRNA29a、miRNA452、miRNA34a及miRNA222表达的影响。结果:β-elemene明显降低耐药乳腺癌细胞株细胞生存率和增殖率,β-elemene显著调节MCF-7/Adr和MCF-7/Doc细胞内miRNAs的表达水平,其中6个miRNAs上调,12个miRNAs下调显著。结论:β-elemene是一种从植物中提取的天然抗肿瘤中药成分,可以逆转乳腺癌细胞的耐药性。证实了中药β-elemene可以通过调控耐药相关miRNA表达的机制抗乳腺癌耐药细胞的耐药性。

Methionine Levels in the Diet for Pregnant Female Rabbits Affects Hair Follicles Development of Baby Rabbits

[ Objective] This experiment was conducted to study methionine level in diet for prepnant famale rabbits on the development of hair follicles of baby rabbits. [ Method] Fifty-four Angora female rabbits with similar body weight, parities and historicaly hair yield were randomly assigned to three groups with eighteen replicates in each group and one rabbit per replicate. Rabbits during pregnancy were fed diets with 0.5!%, 0.71% and 0.91% methionine, respectively. Hair follicle density and diameter in the back skin were determined at 18-day fetus. 26-day fetus, newborn and 2- month-old,and hair fiber diameter at 2-month-old was also determined. [ Result] The results showed as follows： （1） hair follicle density and diameter at each stage,coarse hair fiber diameter at 2-month. old showed a tendency of increasing with dietary methionine level（ P 〉0.05）, fine hair diameter in 0.91% group was significantly larger than that in 0.51% group at 2-month-old （ P 〈 0.05）. （2） With the development of baby rabbit,the ratio of secondary follicle and primary follicle increased gradually, at newborn of rabbits in group 0.51%, 0.71% and 0.91%were 5.84, 5.56 and 5.81, respectively, and at 2-month-old were 8.47, 7.97 and 8.03, respectively. （3） Baby rabbits in group 0.51%, 0.71% and 0.91% gained increases of primary follicle diameter by 0.58, 0.57 and 0.61pm/d from 26-day fetus to birth, and 0.11,0.11 and 0.12 prn/d from birth to 2-month-old, re- spectively; they also gained increases on secondary follicle diameter by 0.0018,0. 012 and 0. 011prn/d from birth to 2-months-old, respectively. [ Conclusion] These results indicate that increase of methionine level in diet for female rabbits during pregnancy is a stimulation factor for the development of hair follicle of baby rabbits, and can improve their hair fiber diameter.

β-elemene联合顺铂抑制Hep-2细胞生长作用的实验研究

目的探讨β-elemene联合顺铂对体外培养的Hep-2细胞(人喉咽癌细胞)生长的抑制作用。方法采用MTT比色法和流式细胞分析技术检测不同组别、不同时间(12、24、48h)对Hep-2细胞的抑制其增殖和诱导其凋亡的作用。结果单纯β-elemene组、单纯顺铂组及β-elemene联合顺铂组,12h时对Hep-2细胞增殖抑制率分别为19.05%、20.12%和22.24%,细胞凋亡率分别为12.21%、17.98%和20.34%;24h时细胞增殖抑制率为40.62%、41.51%和64.65%,细胞凋亡率为25.34%、31.08%和34.62%;48h时细胞增殖抑制率为60.50%、68.23%和71.56%,细胞凋亡率为40.63%、42.96%和63.01%。β-elemene联合顺铂组对Hep-2细胞生长抑制作用和诱导细胞凋亡的作用都明显优于其他各组(P<0.05)。结论β-elemene联合顺铂能够明显抑制Hep-2细胞增殖,诱导Hep-2细胞凋亡,为喉咽癌的化疗提供了一个新的启发。

Inhibitory Effect of β-Elemene Emulsion on Platelet Aggregation and its Mechanism

为阐明β榄香烯的抗血小板作用，本文分别采用比浊法和放免法测定大鼠连续ｉｐ７ｄβ榄香烯乳６．２５～１２．５ｍｇ·ｋｇ－１·ｄ－１后对血小板聚集、血浆ｋｅｔｏＰＧＦ１α和ＴＸＢ２水平的影响。结果表明，本品分别使凝血酶、花生四烯酸和ＡＤＰ诱导血小板最大聚集率下降３８．３％～４２．６％，１４．７％～１９．１％和７．２％～１０．８％，血浆ＴＸＢ２从８８ｎｇ·Ｌ－１下降至７２ｎｇ·Ｌ－１，ｋｅｔｏＰＧＦ１α从１７．５ｎｇ·Ｌ－１增加至２０．９ｎｇ·Ｌ－１。提示ＴＸＢ２降低和ｋｅｔｏＰＧＦ１α值增加是其抑制血小板聚集的作用机制之一。

Comparison of Soleus Muscle Fibers in Rats and Rabbits at Different Stages of Postnatal Development

In the present study,the effects of postnatal development on the number and distribution of skeletal muscle fibers of different types in hind leg of rat and rabbit were studied.The soleus muscles of rats aged 2 days,2,4,6,8,and 10 weeks (body weight 10,32,95,190,280,and 320 g),and rabbits aged 2 days,2,4,8,12,16,20,and 24 weeks (body weight 100,220,400,750,1 200,1 600,2 100,and 2 500 g) were stained with succinic dehydrogenase.With an image analysis system,the X-Y coordinates of fibers were used to analyze the growth-related changes.The results of present study showed that three types of fibers were found in the soleus muscles of rat and rabbit,i.e.,type Ⅰ (slow oxidative),ⅡX (fast oxidative),and ⅡA (fast oxidative glycolytic).The type Ⅰ fibers were present throughout the muscle that had a uniform distribution and tended to increase in number with aging.Type ⅡX fibers were scattered throughout the muscle and decreased markedly in number with aging.Type ⅡA fibers were located at the central and deep regions,and showed a little or no change in number and distribution with aging.While be of age,type ⅡA and ⅡX fibers became restricted to the superficial region.No type ⅡB fibers were detected.Type ⅡA fibers had the largest diameter,type Ⅰ intermediate and type ⅡX the smallest.Mean cross-sectional area of each type fibers of rabbits was larger than that of rats.The present results indicate that the number and distribution of muscle fibers of different types in hind limb of rat and rabbit change with the process of postnatal growth.

Effects of 7-methylxanthine on form-deprivation myopia in pigmented rabbits

AIM: To determine the effect of 7-methylxanthine (7-MX) on the posterior sclera of form-deprivation myopia (FDM) in pigmented rabbits. METHODS: Sixteen pigmented rabbits were monocularly deprived (MD) by suturing the right eyelids after natural eye opening (ten-day old) for a period of 30 days. Two groups of pigmented rabbits were fed either 7-MX (30 mg per kg body weight; n=8) or vehicle control (saline equal volume with 7-MX; n=8). Ocular refractions, axial lengths and body weights were measured at the start and the end of the experiment 30 days later. Electron microscopy was used to measure and determine the collagen fibril diameters in the posterior pole of sclera. RESULTS: In vehicle control MD pigmented rabbits, 30 days of MD produced -1.10D+/- 0.78D of myopia and the axial length increased 0.51mm+/- 0.09mm. In MD pigmented rabbits fed with 7-MX, 30 days of MD induced only -0.21D+/- 0.11D of myopia and the axial length increased 0.07mm+/- 0.10mm. There was significant change in axial length of vehicle control MD pigmented rabbits (13.11mm +/- 0.19mm versus 12.60mm+/- 0.06mm; P =0.03). The changes in refraction and axial length of two MD groups' contalateral eyes during the 30 days were not significantly different (2.75D+/- 0.27D versus 2.75D 0.35D, P>0.05; 12.60mm+/- 0.06mm versus 12.45mm +/- 0.14mm, P >0.05). The weights of the two groups pigmented rabbits had no significant changes (187g+/- 22.1g versus 189g+/- 19.3g, P>0.05). The diameter cif scleral collagen fibers increased in both eyes of 7-MX treated pigmented rabbits. There was significant difference in collagen fibril diameters of inner layer (111.34nm+/- 28.30nm versus 94.80nm +/- 27.52nm, P =0.002) and outer layer (167.92nm +/- 55.82nm versus 144.04nm +/- 47.02nm P =0.016) in the posterior sclera between the myopic eyes of vehicle control MD group and contralateral eyes of 7-MX treated MD group. CONCLUSION: 7-MX appears to prevent FDM in pigmented rabbits by remodeling the posterior sclera.

β-elemene enhances the radiosensitivity of gastric cancer cells by inhibiting Pak1 activation

AIM:To explore the potential of β-elemene as a radiosensitizer for gastric cancer cells and the underlying mechanisms.METHODS:SGC7901,MKN45,MKN28,N87,and AGS human gastric cancer cell lines were used to screen for radioresistant gastric cancer cell lines. A 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium(MTT) assay was used to determine the effects of β-elemene and IPA-3 on cell viability in MKN45 and SGC7901 gastric cancer cell lines. A clonogenic survival assay and annexin V-FITC/PI apoptosis detection assay were used to evaluate cellular radiosensitivity and radiation-induced cell death,respectively. A proteomic method,isobaric tags for relative and absolute quantitation(i TRAQ),was employed to screen the proteins regulated by β-elemene pretreatment prior to ionizing radiation(IR) in SGC7901 gastric cancer cell line. IPA-3 was used as a specific small molecule inhibitor of p21-activated protein kinase 1(Pak1) to target Pak1 signaling. Protein levels of PAK1IP1(p21-activated protein kinase-interacting protein 1),total Pak1(t-Pak1),phospho-Pak1(T423),phospho-ERK1/2( Thr202/Tyr204),and cleaved caspase-3(17 k Da) were assessed by western blotting.RESULTS:MKN45 and SGC7901 gastric cancer cell lines were relatively more resistant to IR. β-elemene pretreatment decreased clonogenic survival following IR in MKN45 and SGC7901 gastric cancer cell lines. Additionally,β-elemene pretreatment prior to IR increased radiation-induced cell death compared with IR alone in MKN45(10.4% ± 0.9% vs 34.8% ± 2.8%,P < 0.05) and SGC7901(11.6% ± 0.9% vs 46.7% ± 5.2%,P < 0.05) human gastric cancer cell lines,respectively,consistent with the level of cleaved caspase-3(17 k Da). Through i TRAQ analysis and western blot validation,we found that β-elemene upregulated PAK1IP1 and downregulated phospho-Pak1(T423) and phosphoERK1/2 in SGC7901 gastric cancer cells. IR increased the level of phospho-Pak1(T423). Pretreatment with β-elemene decreased radiation-induced Pak1 and ERK1/2 phosphorylation. Inhibition of Pak1 using IPA-3 decreased clonogenic survival following IR. In addition,IPA-3 increased radiation-induced cell death in MKN45(13.4% ± 0.3% vs 26.6% ± 1.0%,P < 0.05) and SGC7901(16.0% ± 0.6% vs 37.3% ± 1.7%,P < 0.05) gastric cancer cell lines,respectively,consistent with the level of cleaved caspase-3(17 k Da). Western blotting showed that IPA-3 decreased radiation-induced Pak1 and ERK1/2 phosphorylation.CONCLUSION:This is the first demonstration that β-elemene enhances radiosensitivity of gastric cancer cells,and that the mechanism involves inhibition of Pak1 signaling.

Preparation, characterization, pharmacokinetics and anticancer effects of PEGylated β-elemene liposomes

Objective:This study aimed to develop a new polyethylene glycol(PEG)ylatedβ-elemene liposome(PEG-Lipo-β-E)and evaluate its characterization,pharmacokinetics,antitumor effects and safety in vitro and in vivo.Methods:The liposomes were prepared by ethanol injection and high-pressure micro-jet homogenization.Characterization of the liposomes was conducted,and drug content,entrapment efficiency(EE),in vitro release and stability were studied by ultra-fast liquid chromatography(UFLC)and a liquid surface method.Blood was drawn from rats to establish the pharmacokinetic parameters.The anticancer effect was evaluated in a KU-19-19 bladder cancer xenograft model.Histological analyses were performed to evaluate safety.Results:The PEG-Lipo-β-E showed good stability and was characterized as 83.31±0.181 nm in size,0.279±0.004 in polydispersity index(PDI),-21.4±1.06 mV in zeta potential,6.65±0.02 in pH,5.024±0.107 mg/mL inβ-elemene(β-E)content,and 95.53±1.712%in average EE.The Fourier transform infrared spectroscopy(FTIR)and differential scanning calorimetry(DSC)indicated the formation of PEG-Lipo-β-E.Compared to elemene injection,PEG-Lipo-β-E demonstrated a 1.75-fold decrease in clearance,a 1.62-fold increase in half-life,and a 1.76-fold increase in area under the concentration-time curves(AUCs)from 0 hour to 1.5 hours(P<0.05).PEG-Lipo-β-E also showed an enhanced anticancer effect in vivo.Histological analyses showed that there was no evidence of toxicity to the heart,kidney,liver,lung or spleen.Conclusions:The present study demonstrates PEG-Lipo-β-E as a new formulation with ease of preparation,high EE,good stability,improved bioavailability and antitumor effects.

An ATF24 peptide-functionalized β-elemene-nanostructured lipid carrier combined with cisplatin for bladder cancer treatment

Objective:In this study,we aimed to develop an amino-terminal fragment(ATF)peptide-targeted liposome carryingβ-elemene(ATF24-PEG-Lipo-β-E)for targeted delivery into urokinase plasminogen activator receptor-overexpressing bladder cancer cells combined with cisplatin(DDP)for bladder cancer treatment.Methods:The liposomes were prepared by ethanol injection and high-pressure microjet homogenization.The liposomes were characterized,and the drug content,entrapment efficiency,andin vitro release were studied.The targeting efficiency was investigated using confocal microscopy,ultra-fast liquid chromatography,and an orthotopic bladder cancer model.The effects of ATF24-PEG-Lipo-β-E combined with DDP on cell viability and proliferation were evaluated by a Cell Counting Kit-8(CCK-8)assay,a colony formation assay,and cell apoptosis and cell cycle analyses.The anticancer effects were evaluated in a KU-19-19 bladder cancer xenograft model.Results:ATF24-PEG-Lipo-β-E had small and uniform sizes(~79 nm),high drug loading capacity(~5.24 mg/mL),high entrapment efficiency(98.37±0.95%),and exhibited sustained drug release behavior.ATF24-PEG-Lipo-β-E had better targeting efficiency and higher cytotoxicity than polyethylene glycol(PEG)ylatedβ-elemene liposomes(PEG-Lipo-β-E).DDP,combined with ATF24-PEG-Lipo-β-E,exerted a synergistic effect on cellular apoptosis and cell arrest at the G2/M phase,and these effects were dependent on the caspase-dependent pathway and Cdc25C/Cdc2/cyclin B1 pathways.Furthermore,thein vivo antitumor activity showed that the targeted liposomes effectively inhibited the growth of tumors,using the combined strategy.Conclusions:The present study provided an effective strategy for the targeted delivery ofβ-elemene(β-E)to bladder cancer,and a combined strategy for bladder cancer treatment.