Transformaciones histológicas de la fascia dentada de ratones y ratas en los primeros días posnatales / Histological transformations of fascia dentata of mice and rats in the first postnatal days

Se realizó un estudio descriptivo, observacional y transversal en 160 cortes histológicos de la fascia dentada del hipocampo de ratones BALB/c y ratas Wistar blancas, en el Laboratorio de Investigaciones Biomédicas de la Universidad de Ciencias Médicas de Santiago de Cuba, de septiembre de 2013 a igual mes de 2014, con vistas a determinar las transformaciones histológicas que ocurren en dicha fascia en el segundo y sexto días posnatales. La observación microscópica de estos cortes se realizó empleando del software Image J. La extensión de la fascia al sexto día de vida llegó a ser mayor en los ratones; los máximos incrementos del grosor en ambos tipos de roedores se encontraron en el hilus, y el estrato granuloso fue de menor crecimiento en las ratas. La celularidad en los roedores presentó mayores proporciones en las tres regiones del hilus al segundo día, pero disminuyó en el sexto día, mientras que las zonas relacionadas con el hilus mantuvieron una mayor cantidad de células; sin embargo, el número celular disminuyó en ambas regiones moleculares de la fascia de las ratas.

A descriptive, observational and cross-sectional study was carried out in 160 histological cuts of the hippocampus fascia dentata from mice BALB/c and rats white Wistar, in the Laboratory of Biomedical Investigations from Santiago de Cuba Medical University, from September, 2013 to the same month in 2014, with the aim of determining the histological transformations that take place in this fascia in the second and sixth posnatal days. The microscopic observation of these cuts was carried out using the software Image J. The extension of the fascia at the sixth day of life was larger in the mice; the maximum increases of thickness in both types of rodents were in the hilus, and the granular stratum was of smaller growth in rats. The celularity in the rodents presented larger proportions in the three regions from the hilus at the second day, but it decreased at the sixth day, while the areas related to the hilus maintained a greater quantity of cells; however, the cellular number diminished in both molecular regions of the rats fascia.

Dysfunction of tear film and ultrastructural changes of corneal epithelial cells in castrated male mice / 中华实验眼科杂志

Background Studies indicate that the decline of androgen level affects the secretion function of epithelial cells of meibomian glands and lacrimal glands ,which results in the abnormality of tear quality and quantity.However,whether androgen level regulates the mucin adhesion to ocular surface is still not elucidated.Objective This study was to investigate the tear functional changes and corneal ultrastructural changes in castrated male mice.Methods Fifty-six SPF male BALB/c mice were devided into normal control group, sham group, and orchectomy 1-week group, orchectomy 2-week group, orchectomy 4-week group, orchectomy 6-week group, orchectomy 8-week group according to random number table.Orchectomy was performed in the mice of the orchectomy 1-,2-,4-,6-and 8-week group,and only incision rather than enucleation of testis was carried out in the mice of the sham group.Peripheral venous blood samples were collected during eyeball enucleation in all the mice and serum androgen concentration was detected by radioimmunoassay.Tear secretion level was measured with phenol red thread.Break-up time of tear film (BUT) was tested,and corneal inflorescence staining was scored based on the criterion of Toshida.The corneas of all the mice were collected for ultrastructural examination under the transmission electron microscope.The study complied with the Regulations for the Administration of Affairs Concerning Experimental Animals by State Science and Technology Commission.Results Mouse serum androgen concentration was (573.87±6.74) ng/μl in the normal control group and (579.74-±8.24)ng/μl in the sham group, and there was no significant difference between these two groups (t =1.432, P =0.251) , however, the serum androgen concentration was 0 ng/μl in all castrated mice after orchectomy.The tear secretion levels were (5.26 ± 0.99), (4.89 ± 0.56), (4.62 ± 0.83), (4.28 ± 0.63), (3.36 ± 0.56),(2.60±0.27) and (2.08 ±0.35) mrn/5 minutes in the normal control group, sham group and orchectomy 1-, 2-, 4-,6-and 8-week groups, respectively, and the tear secretion levels were significantly lower in the orchectomy 2-, 4-,6-and 8-week groups than those in the normal control group (all at P＜0.05).The BUT were (68.33±12.86), (62.47± 3.75),(58.67±10.03), (47.17±7.64) ,(39.51±3.39),(32.67±3.88) and (31.00±2.36) seconds in the normal control group, sham group and orchectomy 1-, 2-, 4-, 6-and 8-week groups, respectively, and the BUTs were significantly shorter in the orchectomy 2-,4-, 6-and 8-week groups than those in the normal control group (all at P＜0.05).The fluorescein stain score of corneal epithelium was gradually increased with the lapse of castrated time.The decrease of number,shortening and edema of microvilli as well as separation of desmosomes of corneal epithelial cells were found under the transmission electron microscope in castrated mice.Conclusions Decreasing of serum androgen level, declining of aqueous tears,instability of tear film and damage of corneal epithelial cell ultrastructures occur in castrated male mice, and these manifestations are similar to dry eyes.

Changes of collapsin response mediator protein-2 and its phosphorylated protein levels following optic nerve injury / 中华实验眼科杂志

Background Collapsin response mediator protein-2 (CRMP-2) can promote the growth of axons,but CRMP-2 occurs hyperphosphorylation under the induction of cyclin-dependent kinase-5 (CDK5) after central nervous system injury,which leads to the collapse of the growth cone and hinders the repair of nervous system.Being a central nervous system tissue,whether the expressions of CRMP-2 and its phosphorylated protein (p-CRMP-2) change after optic nerve injury are rarely studied.Objective This study was to investigate the dynamic changes of CRMP-2 and p-CRMP-2 expressions in injured optic nerve tissue.Methods Forty-eight 8-or 9-week-old BALB/c mice were randomly divided into the sham operation group and postoperative 3-,7-and 14-day group.Optic nerves were exposed and clamped at retrobulbar 2 mm for 10 seconds in the right eyes during the surgery in the postoperative 3-,7-and 14-day groups,and the same operation was performed except the clamp of optic nerve in the sham operation group.The optic nerve tissue was obtained from the eyes 3,7 and 14 days after surgery.The relative expression levels of CRMP-2 mRNA and CRMP-2,p-CRMP-2 and CDK5 proteins in the tissue were detected by real-time fluorescence quantitative PCR and Western blot,respectively.The use and care of the experimental animals complied with the Regulation for the Administration of Affair Concerning Experimental Animals of Third Military Medical University.Results No significant differences were found in the expression levels of CRMP-2 mRNA and CRMP-2 protein among the sham operation group and postoperative 3-,7-and 14-day groups (CRMP-2 mRNA:F =2.971,P =0.097;C RMP-2 protein:F=1.202,P =0.370).The relative expression levels of p-CRMP-2 protein in the optical nerve were 0.001±0.000,0.064±0.003,0.136±0.005 and 0.346±0.012,and those of CDK5 protein were 0.440±0.009,0.723±0.011,0.874±0.015 and 0.952±0.019 in the sham operation group and postoperative 3-,7-and 14-day groups respectively,showing statistically significant differences among them (p-CRMP-2:F=445.600,P ＜ 0.001;CDK5:F=186.600,P＜0.001),and the relative expression levels of p-CRMP-2 and CDK5 protein were evidently higher in the optical nerve tissue in the postoperative 3-,7-and 14-day groups than those in the sham operation group (all at P＜0.01).Conclusions There are not significant changes in the expression level of CRMP-2 in the BALB/c mice after optic nerve injury.However,the expression levels of p-CRMP-2 and CDK5 proteins are gradually upregulated as the extending of injured time.

Study on human adipose mesenchymal stem cells differentiating into retinal pigment epithelial-like cells and its in vivo application / 中华实验眼科杂志

Background Stem cell transplantation represents a promising treatment option for patients suffering from degenerative disorders.Accumulating evidences indicate that mesenchymal stem cells (MSCs) are able to differentiate into retinal pigment epithelial (RPE)-like cells.However,MSCs are difficult to obtain.Human adipose mesenchymal stem cells (ADSCs) are proved to have similar properties to MSCs,but relevant study is less.Objective This study was to assess the feasibility of human ADSCs differentiating into RPE-like cells and the safety of its application in vivo.Methods The third generation of human ADSCs were incubated into 6-well plate,and 100 ng/ml epithelial growth factor,50 μ mol/L taurine and 5×10-7 mol/L retinoic acid were added into the medium 12 hours after cultured to induce the cells,and conventional cultured cells were used as the control group.Induced cells were traced with PKH26,and Pan-cytoke ratin (Pan-CK) monoclonal antibody was used to identify the cells under the fluorescence microscope.Induced RPE-like cell suspension of 1 μl was intravetreally injected in the right eyes of 6 BALB/c mice,and equal volume of PBS was used in the same way in another 6 mice.The animals were sacrificed 1 month after injection,and the retinal morphology was examined by histopathology under the optical microscope.The ultrastructure of retinal ganglion cells (RGCs) was examined by the transmission electron microscope.The use and care of the animals complied with Regulations for the Administration of Affair Concerning Experimental Animals by State Science and Technology Commission.Results Cultured human ADSCs grew well with the slender polygone shape.Cell membranes showed the red fluorescence for PKH26 after induced.In addition,Pan-CK was expressed in the cell membranes with the red fluorescence in the induced cells,but the response was absent in the control cells.One month after intravitreal injection,induced cells located on the retinal surface,and the retinal morphology was clear under the optical microscope.No abnormality in RGCs was seen under the transmission electron microscope.Conclusions Human ADSCs can differentiate into RPE-like cells after induction.PKH26 can mark induced cells well.There is no adverse effect of induced cells on retina after intravitreal injection in a short-term duration in mice.

Therapeutic effect of phenylethanoid glycosides on cyclopfosphamide-induced dyszoospermia in mice and its mechannism / 吉林大学学报(医学版)

Objective To explore the therapeutic effect of phenylethanoid glycosides on cyclophosphamide (CTX)-induced dyszoospermia in mice and to preliminary elucidate the mechanisms involved in the process. Methods Phenylethanoid glycoside was extracted by ethanol extraction.Forty male BALB/C mice were randomly divided into control group,model group,low dose of phenylethanoid glycosides group (50 mg· kg-1 )and high dose of phenylethanoid glycosides group (100 mg·kg-1 ).Except control group,the dyszoospermia mouse model was established by peritoneal injection of CTX at the daily dose of 80 mg· kg-1 ,once daily for successive 5 d. After modeling, phenylethanoid glycosides were intragastrically administered at corresponding doses to each phenylethanoid glycosides group.Equal volume of normal saline was given to the mice in control group and model group by gastrogavage.All the medication was performed once daily for successive 30 d.The testis tissue was obtained 24 h after the last intragastric administration.The level of testosterone in the testis tissue homogenate was determined by enzyme linked immunosorbent assay.The sperm counts, the motility rates, and the teratospermia rates in various groups were compared.The morphological changes of the testis tissue were observed using HE staining.Results Compared with control group, the sperm count and the motility rate were decreased, the teratospermia rate was increased,and the testosterone level in the testis tissue homogenate was decreased in model group(P<0.01).Compared with model group,the sperm counts and the motility rates were increased,the teratospermia rates were decreased, and the testosterore levels in the testis tissue homogenate were increased in phenylethanoid glycosides groups (P<0.05 or P<0.01).The histological results showed atrophy and degeneration of seminiferous tuble,thicker seminiferous epithelium and azoospermic lumina in model group;the number of seminiferous epithelial layers was increased and the seminiferous cells orderly arranged, and many sperms were found in the tubules in phenylethanoid glycosides groups.Conclusion Phenylethanoid glycosides has obviously therapeutical effect on CTX-induced dyszoospermia in mice,and its mechanisms might be correlated with recovering the testosterone level.

Effects of combined therapy with thalidomide and glucantime on leishmaniasis induced by Leishmania major in BALB/c mice

For treating Leishmania major infection in BALB/c mice, we used thalidomide in conjunction with glucantime. Groups of mice were challenged with 5 x 10(3) metacyclic promastigotes of L. major subcutaneously. A week after the challenge, drug treatment was started and continued for 12 days. Thalidomide was orally administrated 30 mg/kg/day and glucantime was administrated intraperitoneally (200 mg/kg/day). It was shown that the combined therapy is more effective than single therapies with each one of the drugs since the foot pad swelling in the group of mice received thalidomide and glucantime was significantly decreased (0.9 +/- 0.2 mm) compared to mice treated with either glucantime, thalidomide, or carrier alone (1.2 +/- 0.25, 1.4 +/- 0.3, and 1.7 +/- 0.27 mm, respectively). Cytokine study showed that the effect of thalidomide was not dependent on IL-12; however, it up-regulated IFN-gamma and down-regulated IL-10 production. Conclusively, thalidomide seems promising as a conjunctive therapy with antimony in murine model of visceral leishmaniasis.