Establishment of a rapid method for quality assessment of cardiotonic effect of Shengfupian / 药学学报

Based on the similar structure of adrenaline shared by higenamine (HI), salsolinol (SA) and coryneine (CO), a photochemical colorimetric sensor based on the displacement reaction of o-diphenol hydroxyl group and alizarin red S-phenylboric acid system was constructed to quickly distinguish and identify the cardiac strength of Shengfupian. The results show that the optimal condition of the sensor is: the molar ratio of alizarin red S (ARS) to phenylboric acid (PA) is 1∶3, reaction temperature is 0 ℃; The preparation method of the sample solution is optimized as follows: 2.5 g of Shengfupian powder was taken, 10 times the amount of methanol was added, and 300 W, 40 kHz ultrasound was carried out for 15 min; methodological studies showed that the method had good precision, repeatability and stability. The |△G| value (G is green, |△G| = |G after - G before|) of each sample was obtained by response values determination of 14 batches of Shengfupian. LC-MS/MS was used to determine the contents of three cardiac components in Shengfupian. It was found that the order of the total contents of cardiotonic components was basically consistent with |△G|. Then the correlation was analyzed, and the correlation coefficient R2 was as high as 0.87, which proved the scientificity and accuracy of this method. This study fills the methodological gap of rapid evaluation of the quality of Shengfupian, and provides the key technical support for the high quality and good price of Shengfupian in the market circulation and clinical application.

Effects of atracurium besylate on corneal endothelium of chickens: in vitro study / Avaliação dos efeitos do besilato de atracúrio no endotélio corneano de galinhas: estudo in vitro

The aim of this study was to investigate the acute effects of atracurium besylate on cellular damage in corneal endothelium of chickens. Twenty healthy chicken eyes were assigned to the following groups: Group 1 (G1), experimental group (n=10); and Group 2 (G2), control (n=10). Excised corneoscleral buttons were immediately placed on glass microscopy slides with endothelial region faced up. Corneal endothelium of eyes in G1 were covered with AB (0.2mL, 10mg/mL) for 3 min and then rinsed with balanced salt solution (BSS), while the corneal endothelium of eyes in G2 were covered with BBS for 3 min. Corneas from both groups were stained with alizarin red/trypan blue and visualized by light microscopy. Ten random photographs were taken from each cornea. The area of cellular damage was measured by software in all samples and cell loss of each group was averaged and compared. Endothelial area of denudation and Descemet's membrane exposure were higher in G1 than G2. In conclusion, atracurium besylate induced an acute damage on corneal endothelium of chickens.(AU)'

Objetivou-se avaliar os efeitos agudos do besilato de atracúrio sobre o endotélio corneano de galinhas. Vinte olhos saudáveis de galinhas foram aleatoriamente separados em dois grupos com 10 olhos cada, sendo G1 o grupo controle e G2 o grupo tratamento. Imediatamente após a excisão dos botões corneoesclerais estes foram colocados em lâminas de microscopia de vidro com o lado endotelial voltado para cima. No Grupo 1, o endotélio corneano foi recoberto com 0,2ml de besilato de atracúrio (10mg/ml) durante 3 minutos e depois lavado com solução salina balanceada. No Grupo 2, o endotélio corneano foi recoberto apenas com solução salina balanceada durante 3 min. As córneas de ambos os grupos foram coradas com vermelho de alizarina e azul de tripano e visualizadas com microscópio óptico. Foram obtidas dez fotografias aleatórias de cada amostra. As imagens foram analisadas e com auxílio de um software as áreas com ausência de células endoteliais calculadas. A perda celular endotelial foi significativamente maior no grupo tratamento comparativamente ao grupo controle. Com base nos resultados apresentados foi possível concluir que o besilato de atracúrio induziu dano agudo nas células do endotélio da córnea de galinhas.(AU)

Morphology of endothelial cells from different regions of the cornea of dogs / Morfologia das células endoteliais de diferentes regiões da córnea de cães

ABSTRACT: The aim of this study was to evaluate the morphology of endothelial cells from different areas of the cornea of dogs. Twenty healthy eyes from 10 dogs, females or males, of different ages were studied. Corneal endothelium morphology of superior, inferior, central, nasal and temporal areas was assessed by 0.2% alizarin red staining using an optic microscope. One hundred endothelial cells from each corneal area were analyzed. In all areas of the cornea studied were found endothelial cells with four sides, five sides, six sides and seven sides. There was no significant difference regarding endothelial cell morphology in all corneal regions evaluated. Thus, the morphology of the central cornea area represents the entire endothelial mosaic and may be applied to peripheral areas. Therefore, analysis of the central area is sufficient to estimate the shape of endothelial cells of peripheral areas of healthy dog corneas.

RESUMO: Objetivou-se avaliar a morfologia das células endoteliais de diferentes regiões da córnea de cães. Vinte olhos saudáveis de 10 cães, fêmeas ou machos, de diferentes idades foram estudados. A morfologia do endotélio corneano das regiões superior, inferior, central, nasal e temporal foi avaliada pela coloração vermelho de alizarina 0,2% com microscópio óptico. Foram analisadas 100 células endoteliais de cada região da córnea. Em todas as regiões da córnea estudadas foram encontradas células endoteliais com quatro lados, cinco lados, seis lados e sete lados. Não houve diferença significativa em relação à morfologia de células endoteliais da córnea em todos as regiões estudadas. Assim, a morfologia da região central da córnea representa todo o mosaico endotelial e pode ser aplicada em áreas periféricas. Portanto, a análise da área central é suficiente para estimar a forma das células endoteliais das áreas periféricas de córneas de cães saudáveis.

Alizarin Red-S Protocol for Skeletal Staining during Fetal Period in Rabbit

Background: The main purpose of this study was to give detailed information on the staining protocol of Alizarin Red-S to detect the normal and abnormal skeleton of rabbit fetus. Methods: Eleven (9 females and 2 males) of adult rabbit weighing 3-3.5 kg were used. The female rabbits were left with buck to become pregnant then were classified into a treatment and a control group. The former group received oral doses of 400mg/kg of sodium valproate for 15 days starting from the 6th day after mating until 20th day of pregnancy, while the second received water in the same period. The pregnant rabbit was slaughtered at the 29th day of pregnancy. The live rabbit fetuses were collected. The staining protocol included fixation, dehydration, clearing, staining and preservation. The fetuses were examined under dissecting microscope and photos were taken for documentation. Results: The staining protocol made the rabbit fetuses to be clear enough to see their skeleton through the surrounding tissue. The axial skeleton including skull with mandible, vertebral column, ribs and sternum and the appendicular skeleton including the bones of the fore-and hindlimbs took the stain and became red in color. The macroscopic skeletal disorders of the fetuses of the treatment group were observed. The ossification centres were assessed. Conclusion: This protocol which depended on fixation by 95% ethanol, clearing by 1% potassium hydroxide and staining by 0.001 % Alizarin Red-S was effective in detecting normal and abnormal fetal skeletal morphology

Efficacy of Human Platelet Rich Fibrin Exudate vs Fetal Bovine Serum on Proliferation and Differentiation of Dental Pulp Stem Cells

BACKGROUND AND OBJECTIVES: The imperative role of dental pulp stem cells (DPSCs) in regenerative therapy demands an in-vitro expansion which must deal with the safety and ethical problems associated with fetal bovine serum (FBS). The primary aim of this study was to compare the effects of human platelet rich fibrin (hPRF) exudate Vs FBS on proliferation and osteodifferentiation of human dental pulp stem cells (hDPSCs). The secondary one was to determine the optimum concentration of hPRF exudate inducing hDPSCs proliferation and osteodifferentiation. METHODS: The direct method was used to prepare hPRF exudate. hDPSCs were isolated from impacted mandibular third molars of twelve donors by the outgrowth method. For cell viability and proliferation rate testing, 96 well plates were used and the assay was done in duplicate and the trial repeated four times under the same conditions. Six wells were used to contain 10% FBS, serum free media, 1%, 5%, 10% and 20% concentrations of hPRF exudates, respectively. The proliferation assay was carried out by MTS tetrazolium cell proliferation assay kit and Elisa reader. The study design for osteodifferentiation protocol was exactly as the proliferation one and instead the assay was carried out by alizarin red with Elisa reader. RESULTS: Compared to 10% FBS, 10% hPRF exudate was the optimum concentration for hDPSCs proliferation, while 1% hPRF exudate was the optimum concentration for osteodifferentiation of hDPSCs. CONCLUSIONS: Avoiding the risk of zoonosis which may be occurred with FBS, it is recommended to use 10% hPRF exudate for proliferation and 1% for osteodifferentiation.

Morphology of endothelial cells from different regions of the equine cornea / Morfologia das células do endotélio de diferentes regiões da córnea de equinos

ABSTRACT: The objective of this study was to evaluate the morphology of different regions of the equine cornea using optical microscopy. Both healthy eyes of eight horses, male or female, of different ages were evaluated. Corneas were stained with alizarin red vital dye and subsequently examined and photographed using optical microscopy. Corneal endothelial morphology of central, superior, inferior, temporal and nasal areas was assessed. One hundred endothelial cells from each corneal area were analyzed. The shape of the corneal endothelial cells of each corneal region was analyzed. Statistical data analysis was conducted using the Student's t test. Values of P<0.01 were considered significant. Regarding morphological analysis, no statistically significant differences were reported between the equine corneal regions analyzed. The present research suggested that there are no morphological differences between regions of the equine cornea. The values obtained in any region of the equine cornea can be extrapolated to other regions of the cornea and are representative of the cell morphology present in all regions of the cornea.

RESUMO: O objetivo deste estudo foi o de avaliar a morfologia das diferentes regiões da córnea equina usando microscopia óptica. Foram avaliados ambos os olhos saudáveis ​​de oito equinos, machos ou fêmeas, de diferentes idades. As córneas foram coradas com corante vital vermelho de alizarina, examinadas com microscópio óptico e fotografadas. A morfologia endotélio corneano de áreas centrais, superior, inferior, temporal e nasal foi avaliada. Foram analisadas ​​células endoteliais da córnea de cada área. A forma das células endoteliais da córnea de cada região da córnea foi analisada. Análise estatística dos dados foi realizada por meio do teste t. Valores de P<0.01 foram considerados significativos. Em relação à análise morfológica estatisticamente significativa, não foram encontradas diferenças entre as regiões da córnea equina analisadas. O presente trabalho sugere que não houve diferença entre a morfologia das regiões da córnea de equino. Os valores obtidos em qualquer região da córnea equina podem ser extrapolados para outras regiões da córnea e são representativos da morfologia celular em todas as regiões da córnea.

Protective effect of Cissus quadrangularisLinn. on diabetes induced delayed fetal skeletal ossification.

Background: Delayed fetal skeletal ossification is one of the known complications of maternal diabetes. Objective: The present study was designed to evaluate the protective role of petroleum ether extract of Cissus quadrangularis (PECQ) on diabetes‑induced delayed fetal skeletal ossification. Materials and Methods: Female Wistar rats were rendered diabetic with streptozotocin (STZ, 40 mg/kg, intraperitonial) before mating. After confirmation of pregnancy, the pregnant rats were divided into three groups: normal control group, diabetic control group, and diabetic + CQ group. The diabetic + CQ group pregnant rats were treated with PECQ (500 mg/kg body weight) throughout their gestation period. Immediately after delivery, pups were collected from all three groups and processed for alizarin red S–alcian blue staining in order to examine the pattern of skeletal ossification. Results: Fewer ossification centers and decreased extent of ossification of forelimb and hindlimb bones were observed in the neonatal pups of diabetic control group as compared to those in the normal control group. PECQ pretreatment significantly restored the ossification centers and improved the extent of ossification of forelimb and hindlimb bones in the neonatal pups of diabetic + CQ group as compared to those in the diabetic control group. Conclusions: The results suggested that PECQ treatment is effective against diabetes‑induced delayed fetal skeletal ossification. However, further studies on the isolation and characterization of active constituents of PECQ, which can cross the placental barrier and are responsible for the bone anabolic activity are warranted.

Histopathologic study of lens epithelial cells in different types of cataracts / 中华实验眼科杂志

Background Cataract was directly associated with the damage to the structure and function of lens epithelial cells (LECs).In those patients who suffer from cataracts,morphologic changes of LECs is the most compelling evidence confirming loss of cellular structure and function of LECs.So,learning about the morphological changes of LECs of the different types of cataracts is very important for study on biological behaviors of LECs in different environments or diseases.Objective This study was to evaluate the morphological and ultrastructural changes of the LECs in different types of cataracts.Methods Anterior capsular member from age-related cataracts,diabetic cataracts and high myopia complicated cataract were obtained during the cataract surgery and 15 pieces for each.Trypan blue and alizarin red (TB-AR) stain,haematoxylin and eosin stain were performed in the samples to assess the viability and morphology of LECs.The ultrastructural change of LECs was observed under the transmission electron microscope.The features of the LECs were compared among the different types of cataract.Results TB-AR stain showed that LECs were polygon in shape with the mosaic arrangement and round cell nucleus,and a few dead cells were seen in the samples age-related cataract.In the diabetic samples,LECs largened from swelling with different sizes.More dead cells were found in the high myopia complicated cataract.Haematoxylin and eosin stain exhibited that the anterior capsular membrane presented a homogeneuous membrane,and monolayer LECs attached firmly anterior capsular membrane in the samples of related cataract.Majority of the cells had the intact structure.However,the interspaces between cells and capsular membrane were found in diabetic cataract.Also,smaller LECs were seen in high myopia complicated cataract with the irregular cell morphology.Under the transmission electron microscope,LECs presented the normal shape,intact intercellular tight junction and well attachment between cells and capsular membrane in the samples of the age-related cataract.In the samples of the diabetic cataract,edema of LECs and large intercellular spaces were seen.In addition,the jogged pump and vacuolar degeneration of cytoplasm were revealed in the high myopia complicated cataract.Conclusions The degeneration,necrosis and apoptosis was a common pathological basis of age-related cataract,diabetic cataract and high myopia complicated cataract.However,the damage of LECs was more serious in diabetic cataract and high myopia complicated cataract than that of agerelated cataract.

Evaluation of marking efficiency of different alizarin red S concentrations on body fish structures in Oreochromis niloticus (Perciformes: Cichlidae) juveniles

The use of alizarin red S (ARS) marked tilapias could provide valuable fisheries management information to evaluate fish stocking events and may facilitate aquaculture management practices. As a new technique in fishes, the aim of this study was to compare and evaluate the chemical marks produced in tilapia juveniles by ARS through two treatments: 1) 12 hours of immersion and 2) immersion after osmotic induction. This was analyzed at three concentrations: 50, 75 and 100mg/l, and in three structures: otoliths, fish scales and caudal fin rays of Oreochromis niloticus juveniles. After three culture months 80% of specimens were analyzed and significant differences (p<0.05) in mark intensity were detected between treatments for otoliths and fin rays, but not for fish scales. Significant differences between concentrations were found for the 12h immersion treatment, while no significant differences were detected with osmotic induction. Our results showed that marks appeared at all concentrations, and none of the concentrations produced weak marks. Osmotic induction had a greater mortality than the 12h immersion procedure. After eight culture months the rest of the specimens were analyzed and the mark permanence was observed in all cases. According to the present results we recommend the marking process of 12h immersion treatment at 100mg/L concentration.

El uso de alizarina roja S (ARS) para marcar tilapias podría proporcionar información valiosa para el manejo de su pesquería. Para evaluar pesquerías acuaculturales manejadas con siembras o repoblamientos de peces se comparó y evaluó la marca producida por la alizarina roja S, empleando dos tratamientos: 1) Inmersión en ARS durante 12h; e 2) Inmersión en ARS después de un choque osmótico. El análisis se realizó a tres concentraciones: 50, 75 y 100mg/l y en tres estructuras: otolitos, escamas y radios de la aleta caudal de Oreochromis niloticus. Ochenta por ciento de los ejemplares fueron cultivados durante tres meses y analizados posteriormente. Los resultados mostraron diferencias entre las concentraciones de la marca para el tratamiento de 12h de inmersión mientras que no hubo diferencias entre las concentraciones para el tratamiento con inducción osmótica. Se encontraron diferencias en la intensidad de la marca entre los tratamientos para otolitos y radios de las aletas pero para las escamas no hubo diferencias significativas. Todas las concentraciones produjeron marcas (desde débiles a intensas), sin embargo la concentración de 100mg/l no produjo marcas débiles. El tratamiento por inducción osmótica presentó mayores niveles de mortalidad. Después de ocho meses de cultivo el resto de los ejemplares fueron analizados y se observó la permanencia de las marcas en todos los casos. En vista de lo anterior, para los propósitos de marcaje se recomienda el uso del tratamiento de inmersión por 12h y una concentración de 100mg/l.

Prenatal stress caused by movement restriction induces changes in the appendicular osseous development of CF-1 mice progeny / El estrés prenatal causado por restricción de movimiento induce cambios en el desarrollo óseo apendicular de la progenie de ratones CF-1

Prenatal stress has been associated with alterations in weight and body size, as well as disturbances in the process of developing skeletal ossification, occurring during childbirth and the early stages of life. However, the effect evidence of prenatal stress on bone grow thand development during the gestation period has been low; therefore, it is unknown whether these alterations are associated with potential for growth disorders Because of this, the study aims to determine the short-term effects of prenatal stress on the CF-1mouse bone structure growth in your date of birth. The female mice were divided randomly in two groups: controlled (n=2) and stressed (n=3). The latter was put under stress by means of movement restriction during the last week of gestation. Second, an evaluation of their gestational development was made, obtaining measurements of their weight. Finally, diaphanization with KOH and staining with Alizarin red was used to measure the length of their appendicular bones and their flat pelvic bones, of 53 P0 mice (25 control; 28 stressed during gestation). The stressed mice's body weight (p=0.02) and the length of their appendicular bones (radii, p=0.0011; ulnae, p= 0.0001; humeri, p=0.0001; femorae, p=0.0006; tibiae, p=0.0015) were affected significantly in contrast with the controlled group. On the other hand,there were nosignificant differences inmaternal bodyweightand length ofthe mice pelvic bones (isquium, ilium; p>0.05). The prenatal stress by means of movement restriction alters the osseous appendicular morphology of the CF-1 mouse evaluated at birth...

El estrés prenatal se ha asociado con alteraciones en el peso y el tamaño corporal, así como trastornos en el proceso osificación en el esqueleto en desarrollo, que se producen durante el parto y las primeras etapas de vida. Sin embargo, la evidencia de los efectos del estrés prenatal sobre el crecimiento de los huesos y el desarrollo durante el período de gestación ha sido baja, desconociéndose si estas alteraciones están relacionadas con trastornos del crecimiento. El objetivo fue determinar los efectos a corto plazo del estrés prenatal en la estructura ósea del ratón CF-1 en el día de nacimiento. Las hembras gestantes fueron divididas aleatoriamente en dos grupos: control (n=2) y estresado (n=3). Este último fue puesto bajo estrés por restricción de movimiento durante la última semana de gestación. Evaluando el peso corporal en la progenie al nacer (grupo control n=25; grupo estresado n=28), para posteriormente diafanizar mediante KOH y teñir con alizarina roja, midiendo la longitud de huesos largos apendiculares y huesos planos de pelvis en el plano coronal. El peso de los ratones estresados (p = 0,02) y la longitud de los huesos apendiculares fueron significativamente menores en comparación con el grupo control (radio, p = 0,0011; ulna, p = 0,0001; húmero, p = 0,0001; fémur, p = 0,0006; tibia, p = 0,0015). Por otro lado, no hubo diferencias significativas en el peso corporal de la madre y la longitud de los huesos pélvicos de las crías (isquion, íleon, p> 0,05). El estrés prenatal por restricción de movimiento altera la morfología ósea apendicular del ratón CF-1, evaluados al nacimiento...

In vitro assay for osteoinductive activity of different demineralized freeze-dried bone allograft

PURPOSE: Various bone graft materials have been used for periodontal tissue regeneration. Demineralized freeze-dried bone allograft (DFDBA) is a widely used bone substitute. The current widespread use of DFDBA is based on its potential osteoinductive ability. Due to the lack of verifiable data, the purpose of this study was to assess the osteoinductive activity of different DFDBAs in vitro. METHODS: Sarcoma osteogenic (SaOS-2) cells (human osteoblast-like cells) were exposed to 8 mg/mL and 16 mg/mL concentrations of three commercial types of DFDBA: Osseo+, AlloOss, and Cenobone. The effect of these materials on cell proliferation was determined using the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide assay. The osteoinductive ability was evaluated using alizarin red staining, and the results were confirmed by evaluating osteogenic gene expression using reverse transcription polymerase chain reaction (RT-PCR). RESULTS: In the SaOS-2 cells, an 8 mg/mL concentration of Osseo+ and Cenobone significantly increased cell proliferation in 48 hours after exposure (P<0.001); however, in these two bone materials, the proliferation of cells was significantly decreased after 48 hours of exposure with a 16 mg/mL concentration (P<0.001). The alizarin red staining results demonstrated that the 16 mg/mL concentration of all three tested DFDBA induced complete morphologic differentiation and mineralized nodule production of the SaOS-2 cells. The RT-PCR results revealed osteopontin gene expression at a 16 mg/mL concentration of all three test groups, but not at an 8 mg/mL concentration. CONCLUSIONS: These commercial types of DFDBA are capable of decreasing proliferation and increasing osteogenic differentiation of the SaOS-2 cell line and have osteoinductive activity in vitro.

Petroleum ether extract of Cissus quadrangularis (LINN) stimulates the growth of fetal bone during intra uterine developmental period: a morphometric analysis

OBJECTIVE: The aim of the present study was to analyze the effect Cissus quadrangularis plant petroleum ether extract on the development of long bones during the intra-uterine developmental stage in rats. METHODS: Pregnant rats (n=12) were randomly assigned into either a control group (n=6) or a Cissus quadrangularis treatment (n=6) group. Pregnant rats in the Cissus quadrangularis group were treated with Cissus quadrangularis petroleum ether extract at a dose of 500 mg/kg body weight from gestation day 9 until delivery. The animals in the control group received an equal volume of saline. Newborn pups were collected from both groups for alizarin red S - alcian blue staining to differentiate ossified and unossified cartilage. The ossified cartilage (bone) was morphometrically analyzed using Scion image software. RESULTS: Morphometric analysis revealed that the percentage of the total length of ossified cartilage (bone) in pups born to treated dams was significantly higher (P<0.001- -0.0001) than that of the control group. CONCLUSION: The results of the present study suggest that maternal administration of Cissus quadrangularis petroleum ether extract during pregnancy can stimulate the development of fetal bone growth during the intra-uterine developmental period.

Confocal laser scanning microscopy study of dentinal tubules in dental caries stained with alizarin red

Dentin morphology and the lesion found in dental caries have been studied for many years. It was first observed under optical microscopy, and later using electron microscopy. Confocal laser scanning microscopy (CLSM) applied with several fluorescent dyes such as alizarin red to see normal dentinal tubules. However, as far as authors aware, the CLSM studies of dentinal tubules in human caries using alizarin red is rare. The aim of this study is to examine histopathological and morphological changes in dentinal tubules of dentin caries stained with alizarin red using CLSM. Fifteen extracted carious teeth (premolar or molar) was collected and fixed in neutral formalin solution buffered with phosphate buffer, rinsed and stored in calcium free phosphate buffer saline (PBS) at 4°C. The specimens were dehydrated and embedded in resin. Longitudinal or cross sections were cut and polished and then stained with alizarin red S (100 μg/ml) in 0.5 M HCl solution for 24-48 hour at 37°C. After dehydration specimens were mounted on glass slide and examined under CLSM using epi-flourescent mode or transmission light mode with wave length of 512 nm. The images of dentinal tubules were taken serially and optimum images of three-dimensional structures were reconstructed using software of CLSM. Histopathological changes of dentinal tubules in human caries showed area of demineralized dentin, translucent zone, and normal area. The dentinal tubules were thin and had numerous branches. In conclusion, confocal microscopy revealed Study shows that confocal microscopy revealed histopathological changes in dentinal tubules affected by carious lesions.

Experimental study on rat corneal endothelial damage and repair after acid burn / 第三军医大学学报

Objective To study the effect of acid burn on rat corneal endothelial damage and its repair.Methods The model of mild,moderate and severe acid burn in cornea was established in 90 Long-Evans rats(n=30 for each degree of acid burn) with 4 rats served as normal controls.Corneal edema,opacity and the healing of epithelium were observed with a slit lamp microscope.The corneal endothelium from half rats of each degree acid burn was stained with trypan blue and alizarin red,and that from the other half with hematoxylin and eosin(HE) immediately,30 min,1,3,7,14,21,28 d after cautery.The endothelial damage and its repair were observed and the number of the leucocytes in cornea was calculated under light microscope.Results No damage was seen in the corneal endothelial cells of mild,moderate acid burn,only a few leucocytes could be seen in the cornea,and the healing of epithelium,the regression of edema were within 7 d.The endothelial cells necrosed and shedded in the wound area by severe acid burn.The endothelial cells in the margin were deformed,elongated and slided to the damaged area during repair,and a great number of leucocytes infiltrated,the healing of corneal epithelium was delayed,the regression of edema was up to 28 d.Conclusion The corneal endothelial cells were damaged in severe acid burn.The secondary inflammatory reaction took part in the damage of corneal endothelial cells after severe acid burn.The damage of endothelial cells delayed the healing of corneal ulcer and edema.

Corneal Endothelial Damage after Deep Excimer Laser Ablation

To investigate the effect of deep excimer laser ablation on the corneal endothelium, excimer laser photo refractive keratectomy(PRK) with different ablation depths were performed to obtain various residual corneal thickness(range: 90-250 micrometer) in white rabbit corneas(N=50). Corneal endothelium was stained with Alizarin red S(pH 4.2) for 2 minutes three days after excimer laser ablation and was analyzed morphometrically using digitizer the morphometric parameters according to residual corneal thickness. In the corneas of residual corneal thickness(RT) over 200 micrometer and untreated controls(N=6), endothelial damages were rarely seen. With the decrease of residual corneal thickness, hexagonality and shape factor decreased, and coefficient of variation of the cell area(CV) increased(p<0.01). In corneas of RT less than 200 micrometer, endothelial damages were found and become more severe in corneas of RT between 90~149 micrometer. Hexagonal shaped cells were rarely observed, and the shapes of most cells were changed. Deep excimer laser PRK might affect corneal endothelial cells if the RT is less than 200 micrometer, and these findings suggest that care is recommended when doing deep excimer laser corneal ablation, especially excimer laser assisted in situ keratomileusis.

Subepidermal Calcified Nodule

Subepidermal calcified nodule(SCN) is a form of calcinosis cutis which is usually present as a single small, raised, hard nodule with verrucous surface. A 13-year-old girl presented with a 10-month history of a ricegrain-sized, hard, yellowish white papule on both upper eyelids. The lesions were asymptomatic and had increased slowly in size. An excisional biopsy specimen of the left upper eyelid showed acanthosis and narrow pointed rete ridges of the epidermis, and closely aggregated deposition of basophilic material in the uppermost dermis. The material in the dermis did not stain with von Kossa. It was confirmed as calcium deposition by staining with alizarin red S which is far more specific for calcium than the von Kossa stain.

Morphometric Analysis of the Rabbit Corneal Endothelium Specular Microscopy Versus Alizarin Red S Staining

Analysis of endothelial morphology by computer assisted digitizer and image analysis program provides very useful indices of cell shape and size which appear to correlate to the monolayer's functional status. To study the influences of the Alizarin red S staining, which is commonly used in animal experiments of corneal endothelium and vital staining, on the morphologic characteristics of corneal endothelium, morphometric data(density, area, coefficient of variation, perimeter, shape factor, hexagonality, lengths) obtained by specular microscopy of the endothelium are compared to data obtained by Alizarin red S staining of the endothelium of the excised cornea. Mean endothelial cell area was measured as 389.58 +/- 37.14 micrometer2, density was 2588 +/- 251 cells/mm2. The corresponding values measured after Alizarin red S staining, cell area was 407.42 +/- 45.3 micrometer2 and density was 2484 +/- 294 cells/mm2. But no significant differences were noted in comparing all morphometric data obtained by staining to that obtained from specular microscopy. Therefore, Alizarin red S staining combined with cell morphometric analysis could provide valuable data in a cornea which lacks clarity limits or precludes specular microscopy.

COMPOUND STAINING METHOD SHOWING OSSIFIED SKELETAL STRUCTURES AND CARTILAGE IN FETAL, NEONATAL AND ADULT RAT / 解剖学报

Objective To compare compound staining of Alizarin red S and Alcian blue showing ossified skeletal structures and cartilage in fetal,neonatal and adult rat. Methods All specimens were skinned and fat tissue removed completely and eviscerated,then fixed in 95% ethanol;Acetone was used remove fats; 0.1% Alizarin red S and 0.15%-0.3% Alcian blue were used in ossified bone and cartilage;Differentiation and clearing in 0.5%-2% aqueous KOH. Results Ossified bones showed rose red;cartilage showed blue;the others were transparent.Conclusion The best results of Alizarin red S and Alician blue compound staining could be obtained in all rat skinned,eviscerated,and removed the adipose tissues.