Assessing the biocompatibility of bovine tendon scaffold, a step forward in tendon tissue engineering.

Tendon is a collagen-enriched, tough, and intricately arranged connective tissue that connects muscle to the bone and transmits forces, resulting in joint movement. High mechanical demands can affect normal tissues and may lead to severe disorders, which usually require replacement of the damaged tendon. In recent decades, various decellularization methods have been studied for tissue engineering applications. One of the major challenges in tendon decellularization is preservation of the tendon extracellular matrix (ECM) architecture to maintain natural tissue characteristics. The aim of the present study was to create a decellularized bovine Achilles tendon scaffold to investigate its cytocompatibility with seeded hAd-MSCs (human adipose derived-mesenchymal stem cells) and blastema tissue in vitro. Here, we describe a reliable procedure to decellularize bovine Achilles tendon using a combination of physical and chemical treatments including repetitive freeze-thaw cycles and the ionic detergent SDS, respectively. The decellularization effectiveness and cytocompatibility of the tendon scaffolds were verified by histological studies and scanning electron microscopy for up to 30 days after culture. Histological studies revealed hAd-MSC attachment and penetration into the scaffolds at 5, 10, 15 and 20 days of culture. However, a decrease in cell number was observed on days 25 and 30 after culture in vitro. Moreover, migration of the blastema tissue cells into the scaffold were shown at 10 to 25 days post culture, however, destruction of the scaffolds and reduction in cell number were observed on 30th day after culture. Our results suggest that this decellularization protocol is an effective and biocompatible procedure which supports the maintenance and growth of both hAd-MSCs and blastema cells, and thus might be promising for tendon tissue engineering.

Decellularized bovine aorta as a promising 3D elastin scaffold for vascular tissue engineering applications.

Aim: To evaluate the suitability of using aorta elastin scaffold, in combination with human adipose-derived mesenchymal stem cells (hAd-MSCs), as an approach for cardiovascular tissue engineering. Materials & Methods: Human adipose-derived MSCs were seeded on elastin samples of decellularized bovine aorta. The samples were cultured in vitro to investigate the inductive effects of this scaffold on the cells. The results were evaluated using histological, and immunohistochemical methods, as well as MTT assay, DNA content, reverse transcription-PCR and scanning electron microscopy. Results: Histological staining and DNA content confirmed the efficacy of decellularization procedure (82% DNA removal). MTT assay showed the construct's ability to support cell viability and proliferation. Cell differentiation was confirmed by reverse transcription-PCR and positive immunohistochemistry for alfa smooth muscle actin and von Willebrand. Conclusion: The prepared aortic elastin samples act as a potential scaffold, in combination with MSCs, for applications in cardiovascular tissue engineering. Further experiments in animal models are required to confirm this.

A comparison study of different decellularization treatments on bovine articular cartilage.

Previous researches have emphasized on suitability of decellularized tissues for regenerative applications. The decellularization of cartilage tissue has always been a challenge as the final product must be balanced in both immunogenic residue and mechanical properties. This study was designed to compare and optimize the efficacy of the most common chemical decellularization treatments on articular cartilage. Freeze/thaw cycles, trypsin, ethylenediaminetetraacetic acid (EDTA), sodium dodecyl sulfate (SDS), and Triton-X 100 were used at various concentrations and time durations for decellularization of bovine distal femoral joint cartilage samples. Histological staining, scanning electron microscopy, DNA quantification, compressive strength test, and Fourier-transform infrared spectroscopy were performed for evaluation of the decellularized cartilage samples. Treatment with 0.05% trypsin/EDTA for 1 day followed by 3% SDS for 2 days and 3% Triton X-100 for another 2 days resulted in significant reduction in DNA content and simultaneous maintenance of mechanical properties. Seeding the human adipose-derived stem cells onto the decellularized cartilage confirmed its biocompatibility. According to our findings, an optimized physiochemical decellularization method can yield in a nonimmunogenic biomechanically compatible decellularized tissue for cartilage regeneration application.

Use of cerium oxide nanoparticles: a good candidate to improve skin tissue engineering.

Today advancements in nanotechnology have made extensive progress in tissue engineering. Application of cerium oxide nanoparticles (CeO2) has improved regenerative medicine due to their antioxidant properties. In this study, nanoparticles were used to increase the efficacy of skin substitutes. Human skin samples were decellularized using four methods and studied via histological stainings and DNA content analyses. Then CeO2 dispersing and its stability were investigated. The prepared acellular dermal matrices (ADMs) were immersed in CeO2 suspension and their effects were evaluated on growth of cultured human adipose derived-mesenchymal stem cells (hAd-MSCs) using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and histological methods. Moreover, their antioxidant properties were assessed based on DPPH degradation. Changes in the collagen contents of the scaffolds containing cells and CeO2 were also determined by electron microscopy and their tensile strength was compared to ADM. Our results indicated that use of trypsin/NaOH protocol resulted in most efficient cell removal while maintaining extracellular matrix (ECM) architecture. Among different dispersal methods, the approach using Dulbecco's modified Eagle's medium (DMEM), wetting with fetal bovine serum (FBS) and ultrasonic bath resulted in the best stability. Furthermore, it was shown that CeO2 not only had no toxicity on the cells, but also increased the growth and survival of hAd-MSCs by about 27%, improved free radical scavenging, as well as the amount of collagen and tensile strength of the scaffolds containing nanoparticles compared to the ADM. It can be concluded that the combination of ADM/CeO2/hAd-MSCs could be a step forward in skin tissue engineering.

Development of a demineralized and decellularized human epiphyseal bone scaffold for tissue engineering: A histological study.

Extracellular matrix (ECM) controls cellular behaviors such as proliferation, migration, and differentiation. The decellularized matrix of mammalian tissues has been used as a scaffold for the repair and reconstruction of tissue defects. In this study, for the first time demineralized and decellularized human epiphyseal bone matrix was used as a scaffold for a bone generation. Human epiphyseal bone was demineralized by hydrochloric acid and then decellularized by three methods of physical (slow freezing and snap freeze-thaw), enzymatic (trypsin 0.25%, 18 h) and chemical sodium dodecyl sulfate (SDS),)2.5%, 26 h). The scaffolds were cultured with rat adherent bone marrow cells (RABMC). Then, the histological studies were performed on days 7, 14, 21, and 28 of the culture to observe the distribution and morphology of cells. Bone formation was also investigated using Alizarin red staining. The results of this study indicated that RABMC migrated, proliferated and separated by forming lacuna in this three-dimensional bone scaffold. In addition, the Alizarin red staining indicated the calcium deposition on the scaffold in both bone differentiation and standard culture medium. The natural characteristic of the present bone scaffold, its cell adhesion features and capability to induce bone mineralization, even in the standard culture medium, provides a potentially optimal bone scaffold for bone tissue engineering.

Supportive properties of basement membrane layer of human amniotic membrane enable development of tissue engineering applications.

Human amniotic membrane (HAM) has been widely used as a natural scaffold in tissue engineering due to many of its unique biological properties such as providing growth factors, cytokines and tissue inhibitors of metalloproteinases. This study aimed at finding the most suitable and supportive layer of HAM as a delivery system for autologous or allogeneic cell transplantation. Three different layers of HAM were examined including basement membrane, epithelial and stromal layers. In order to prepare the basement membrane, de-epithelialization was performed using 0.5 M NaOH and its efficiency was investigated by histological stainings, DNA quantification, biomechanical testing and electron microscopy. Adipose-derived stromal cells (ASCs) and a human immortalized keratinocyte cell line (HaCaT) were seeded on the three different layers of HAM and cultured for 3 weeks. The potential of the three different layers of HAM to support the attachment and viability of cells were then monitored by histology, electron microscopy and (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Moreover, mechanical strengths of the basement membrane were assessed before and after cell culture. The results indicated that the integrity of extra cellular matrix (ECM) components was preserved after de-epithelialization and resulted in producing an intact basement amniotic membrane (BAM). Moreover, all three layers of HAM could support the attachment and proliferation of cells with no visible cytotoxic effects. However, the growth and viability of both cell types on the BAM were significantly higher than the other two layers. We conclude that growth stimulating effectors of BAM and its increased mechanical strength after culturing of ASCs, besides lack of immunogenicity make it an ideal model for delivering allogeneic cells and tissue engineering applications.

Effects of Neonatal Exposure to Zearalenone on Puberty Timing, Hypothalamic Nuclei of AVPV and ARC, and Reproductive Functions in Female Mice.

It is now established that mycoestrogen zearalenone (ZEN) disrupts reproductive physiology, but the specific mechanisms by which this occurs remain unknown, especially in brain. Growing evidence suggests that populations of estradiol (E2)-sensitive neurons in anteroventral periventricular (AVPV) and arcuate (ARC) nuclei, especially kisspeptin neurons, play a pivotal role in the timing of puberty onset, ovulation, and normal reproduction. The present study was conducted to find whether the ZEN can cause estrogen-like actions during the critical period of neonatal differentiation. In this study, we compared the effect of neonatal exposure to sesame oil, E2 benzoate (EB, 20 µg/kg body weight [bw]), and 3 various doses: 0.2, 1, and 2 mg/kg bw of ZEN (0.2, 1, and 2 ZEN) on the onset of puberty and estrus cyclicity as well as ovarian follicular profile, kisspeptin expression, and neuronal density in AVPV and ARC hypothalamic nuclei and E2 and luteinizing hormone (LH) levels on postnatal day 70. Control mice received no treatment. Vaginal opening was significantly advanced by EB and 2 ZEN. Disrupted estrus cycles and decreased follicular profiles were observed in EB, 1 ZEN, and 2 ZEN animals. In addition, EB, 1 ZEN, and 2 ZEN reduced the expression of kisspeptin and neuronal density of AVPV and ARC nuclei and caused a decrease in the LH and an increase in E2 plasma levels. Taken together, our observations provide physiological evidence that neonatal exposure to ZEN exerts estrogen-like actions in the estrogen-sensitive hypothalamic AVPV and ARC nuclei, controlling reproductive functions in adult female mice.

Comparative Dermatoglyphic Study between Autistic Patients and Normal People in Iran.

Autism is a neurodevelopmental disorder originating from early childhood; nevertheless, its diagnosis is in older ages. In addition to heredity, environmental factors are also of great significance in the etiology of the disease. Dermatoglyphic patterns, albeit varied, remain stable for a lifetime and yield a large number of patterns upon examination. Studies have shown a significant association between dermatoglyphics and some diseases, especially genetic ones. We compared fingerprints between patients with autism and normal individuals in a Fars population living in Khorasan-Razavi Province, Iran, in 2015. The right and left hand fingerprints of 104 autistic individuals (case group; age range=5-15 y) were collected using a fingerprint scanner. The same process was performed for 102 healthy individuals, in the age range of 6 to 25 years. All dermatoglyphic patterns and ridge counts were determined. The data were analyzed using the Mann-Whitney nonparametric test and binomial distribution. There was a significant difference in the distribution of the dermatoglyphic patterns on the right and left thumbs and the index fingers between the case and control groups (P<0.05). The patients had a significantly higher count of loops on their right and left thumbs and their index fingers. A significant decrease in ridge counts for the right and left thumbs and the index fingers was observed in the patients compared to the controls. The results suggested that the patterns were associated with the risk of autism. The patterns may be drawn upon as biometric parameters in the screening of children with autism.

Oestrogenic action of neonatal tamoxifen on the hypothalamus and reproductive system in female mice.

Tamoxifen, a selective oestrogen receptor modulator, is widely used for both the treatment and prevention of breast cancer in women; however, it is known to have adverse effects in the female reproductive system. Growing evidence suggests that oestrogen-sensitive neuron populations of the anteroventral periventricular (AVPV) nucleus and arcuate (ARC) nucleus, especially kisspeptin neurons, play a pivotal role in the timing of puberty onset and reproductive function. The aim of the present study was to evaluate whether neonatal exposure to tamoxifen affects oestrogenic actions in the brain and reproductive function in mice. On 1 to 5 postnatal days, female pups were injected subcutaneously with sesame oil (sham), oestradiol benzoate (EB; 20 µg kg-1), tamoxifen (0.4 mg kg-1) or EB+tamoxifen. Control mice received no treatment. Mice in the EB, tamoxifen and tamoxifen+EB groups exhibited advanced vaginal opening, disrupted oestrous cycles and a decreased follicular pool. Conversely, in these groups, there was a reduction in kisspeptin (Kiss1) mRNA expression, the neuronal density of AVPV and ARC nuclei and LH and oestradiol concentrations in the serum. The results of the present study confirm oestrogenic actions of tamoxifen in the brain and reproductive system. In addition, we show, for the first time, that tamoxifen has oestrogenic effects on the oestrogen-sensitive hypothalamic AVPV and ARC nuclei controlling the reproductive axis in female mice.

Evaluation of Antimicrobial and Healing Activities of Frog Skin on Guinea Pigs Wounds.

BACKGROUND: Frog skin secretions have potentials against a wide spectrum of bacteria. Also, frog skin compositions have healing properties. OBJECTIVES: The aim of this study was to investigate the antibacterial potentials along with healing properties of frog skin Rana ridibunda, a species which thoroughly lives in Iran marshes, as a biological dressing on wounds. MATERIALS AND METHODS: In this study, excisional wounds, dressed with frog skin, were compared between experimental and control groups of guinea pigs. In the experimental groups, wounds were dressed with the dermal (FS) and epidermal (RFS) sides of fresh frog R. ridibunda skin, while only usual cotton gauze covered the wounds of the control group. Furthermore, microbial samples were taken on different days (0, 3, 5, and 7 days post injury) to count the number of the colony-forming units. Additionally, the microbial penetration test was performed on frog skin and then the progression of wound closure was evaluated. RESULTS: In the microbial studies, the bacterial load considerably declined in the wounds treated with FS and RFS compared with the control wounds. On day 7 post injury, the numbers of the colony-forming units for the FS, RFS, and control groups were 6.75, 105, and 375, respectively. In the penetration test, fresh frog skin showed to be a bacterial resistant dressing. The results revealed that the rate of wound closure in the experimental groups significantly was accelerated in comparison with that in the control group. CONCLUSIONS: Our results demonstrated the antimicrobial properties of frog skin as a wound dressing, which has antimicrobial effects per se. This biological dressing shows promise as an effective biological wound dressing insofar as not only is it capable of resisting microbes and accelerating wound healing but also it is cost-effective and easy to use.

Use of an in vitro model in tissue engineering to study wound repair and differentiation of blastema tissue from rabbit pinna.

Rabbit ear wound repair is an accepted model for studies of tissue regeneration, leading to scar less wound repair. It is believed that a specific tissue, blastema, is responsible for such interesting capacity of tissue regeneration. To test this idea further and to elucidate the cellular events happening during the ear wound repair, we designed some controlled experiments in vitro. Small pieces of the ear were punched and washed immediately with normal saline. The tissues were then cultured in the Dulbecco's Modified Eagle(')s Medium, supplemented with fetal bovine serum in control group. As a treatment vitamin A and C was used to evaluate the differentiation potency of the tissue. These tissues were fixed, sectioned, stained, and microscopically studied. Micrographs of electron microscopy provided evidences revealing dedifferentiation of certain cells inside the punched tissues after incubation in tissue culture medium. The histological studies revealed that cells of the tissue (i) can undergo cellular proliferation, (ii) differentiate to epithelial, condrogenic, and osteogenic tissues, and (iii) regenerate the wounds. These results could be used for interpretation of the possible events happening during tissue engineering and wound repair in vitro. An important goal of this study is to create a tissue engineering and tissue banking model, so that in the future it could be used in further blastema tissue studies at different levels.

In vitro histological investigation of interactions between rat decellularized large intestine scaffold and human adipose derived mesenchymal stem cells.

The aim of this study was to investigate the interactions between rat intestine decellularized scaffold and human adipose derived mesenchymal stem cells. Rat large intestine was dissected in fragments and decellularized by physicochemical methods. The scaffolds were loaded by human adipose derived mesenchymal stem cells expressing green fluorescent protein. Microscopic sections were prepared from the scaffolds after two weeks of culture with stem cells and studied by histological methods. The interactions of scaffolds with MSCs were also studied by electron microscopy. Histological and electron microscopy studies revealed human mesenchymal stem cell adhesion, migration, division and maintenance during the 14 days of culture in vitro. According to the results, scaffolds prepared from rat intestine matrix could be a suitable scaffold for studying in vitro cell behaviors such as division, migration and attachment. These various behaviors of cultured cells might be due to inductive effects of the extracellular matrix derived scaffold. However, more investigations are required to discover the exact effects of this scaffold and its interactions with mesenchymal stem cells.

Mesenchymal stem cells can survive on the extracellular matrix-derived decellularized bovine articular cartilage scaffold.

OBJECTIVE S: The scarcity of articular cartilage defect to repair due to absence of blood vessels and tissue engineering is one of the promising approaches for cartilage regeneration. The objective of this study was to prepare an extracellular matrix derived decellularized bovine articular cartilage scaffold and investigate its interactions with seeded rat bone marrow mesenchymal stem cells (BM-MSCs). MATERIALS AND METHODS: Bovine articular cartilage that was cut into pieces with 2 mm thickness, were decellularized by combination of physical and chemical methods including snap freeze-thaw and treatment with sodium dodecyl sulfate (SDS). The scaffolds were then seeded with 1, 1'-dioctadecyl-3, 3, 3', 3'-tetramethylindocarbocyanine perchlorate (DiI) labeled BM-MSCs and cultured for up to two weeks. RESULTS: Histological studies of decellularized bovine articular cartilage showed that using 5 cycles of snap freeze-thaw in liquid nitrogen and treatment with 2.5% SDS for 4 hr led to the best decellularization, while preserving the articular cartilage structure. Adherence and penetration of seeded BM-MSCs on to the scaffold were displayed by histological and florescence examinations and also confirmed by electron microscopy. CONCLUSION: ECM-derived decellularized articular cartilage scaffold provides a suitable environment to support adhesion and maintenance of cultured BM-MSCs and could be applied to investigate cellular behaviors in this system and may also be useful for studies of cartilage tissue engineering.

The effect of maternal nicotine on basement membrane collagen IV of brain microvessels changes in neonatal Balb/C mice.

BACKGROUND: Nicotine can pass through placental blood barrier and accumulate in the developing organs of fetus. Also, entering the breast milk, nicotine can have an effect on the neonates. Investigations have showed that collagen IV is one of the most important micro vessels basement membrane components. OBJECTIVE: In this study, the effect of maternal nicotine exposure in pre and postnatal periods on collagen IV in microvessels of neonatal Balb/C mice brain cortex was studied by immunohistochemistry technique. MATERIALS AND METHODS: 24 pregnant Balb/C mice were divided in to 4 groups (6 mice in each group): two experimental and 2 control groups. The mothers in the 1(st) experimental group were injected 3 mg/kg nicotine intrapritoneally from the 5(th) day of pregnancy to parturition daily and in 2(nd) experimental group the same procedure was repeated to the 10(th) day after parturition (lactation). The control groups received the same volume of normal saline during the same time. 10 days after delivery, the brain tissues of newborns were isolated. Then, prepared blocks from fixed brain were cut serially for immunohistochemical assay. RESULTS: The findings of the present study indicated that collagen IV reaction in microvessels basement membrane in the first experimental group increased significantly compared to the first control group (p=0.002). In addition, collagen IV reaction in microvessels basement membrane in the 2(nd) experimental group increased significantly compared to the 2(nd) control group (p=0.002). However, no significant difference was observed between the two experimental groups. CONCLUSION: These results suggested that maternal nicotine exposure during prenatal period may increase basement membrane collagen IV expression. Also, nicotine increases in maternal breast milk has no effect on basement membrane collagen IV expression.

Pharmaceutical application of frog skin on full-thickness skin wound healing in mice.

CONTEXT: It has been proved that fresh frog skin is efficient in the wound healing process. OBJECTIVE: The purpose of study is to introduce a formulation of frog skin powder for evaluation of wound repair where fresh frog skin is not available. MATERIALS AND METHODS: Rana ridibunda (Ranidae) skins were lyophilized, and a powder was prepared. The powder (0.0005 g) was then mixed with ointment (0.0065 g) for treating each wound. Formulation was used on full-thickness wounds on mice (FO group) and compared to positive and negative controls. In order to study the wound healing process, wound contraction, inflammation, number of fibroblast cells, neovascularization and collagen density were evaluated on days 2, 4 and 6 following the injury. Moreover, CFU measurement was performed for the evaluation of wound contamination. RESULTS: Acceleration in wound contraction in the FO group compared to control groups was significant (p < 0.001) on days 4 and 6. Results showed that FO treatment considerably decreased inflammatory cells during the study. On day 4, FO treatment was significantly effective in increasing the number of fibroblast cells and collagen density (p < 0.01 and p < 0.05, respectively). On day 6 the number of fibroblast cells (p < 0.001), collagen density (p < 0.05) and neovascularization (p < 0.05), were higher in the FO group than the control groups. Results of CFU measurement demonstrated significant reduction of wound contamination in FO treated wounds on days 2 (p < 0.05) and 4 (p < 0.01). DISCUSSION AND CONCLUSION: Our findings indicated that the pharmaceutical form of frog skin used in this study has considerable healing and antibacterial effects on wounds.

The effect of maternal thyroid disorders (hypothyroidism and hyperthyroidism) during pregnancy and lactation on skin development in wistar rat newborns.

OBJECTIVE(S): Previous studies have shown that thyroid hormones are necessary for normal development of many organs and because of the importance of skin as the largest and the most important organ in human body protection in spite of external environment, the study of thyroid hormones effects on skin development is considerable. In this survey we have tried to study the effects of maternal hypothyroidism on skin development in fetus during pregnancy and lactation by immunohistochemistry technique. MATERIALS AND METHODS: Rats were divided into 4 groups, hypothyroids, hyperthyroids, hypothyroids are treated with levothyroxin and a control group. The rat mothers were exposed to PTU with 50 mg/lit dosage and levothyroxin with 1 mg/lit dosage and PTU and levothyroxin simultaneously and with the same dosage respectively in hypothyroid, hyperthyroid and treated hypothyroids with levothyroxin groups. After 14 days, blood sample was taken from mothers, and if thyroid hormones level had change well, mating was allowed. After pregnancy and delivery, 1th day dorsal skin (as the sample for pregnancy assay) and 10th day skin (as for lactation assay) was used for immunohystochemical and morphometric studies. RESULTS: In this study it was observed that maternal hypothyroidism during pregnancy and lactation causes significant increase in laminin expression, in most areas of skin, and maternal hyperthyroidism during pregnancy and lactation causes significant decrease in laminin expression. Also significant decrease was observed in hair follicles number and epidermis thickness in hypothyroidism groups. CONCLUSION: This study showed maternal hypothyroidism causes significant decrease in epidermis thickness and hair follicles number and it causes less hair in fetus. Also maternal hypothyroidism causes large changes in laminin expression in different parts of skin. At the same time,maternal hyperthyroidism causes opposite results. In fact, thyroid hormones regulate laminin expression negatively which means increase in thyroid hormone level, decreases laminin expression. So changes in thyroid hormones level can influence skin development significantly.

Three-dimensional scaffold from decellularized human gingiva for cell cultures: glycoconjugates and cell behavior.

OBJECTIVE: We studied both the presence of some carbohydrate compounds in a threedimensional (3D) matrix harvested from human gingiva and the cell behavior in this matrix. MATERIALS AND METHODS: In this experimental research, in order to prepare 3D scaffolds, human palatal gingival biopsies were harvested and physically decellularized by freezethawing and sodium dodecyl sulfate (SDS). The scaffolds were placed within the rings of blastema tissues obtained from a pinna rabbit, in vitro. We evaluated the presence of glycoconjugatesand cellular behavior according to histological, histochemical and spectrophotometry techniques at one, two and three weeks after culture. One-way analysis of variance (ANOVA)comparedthe groups. RESULTS: Extracellular matrix (ECM) remained after decellularization of tissue with 1% SDS. Glycoconjugate contents decreased meaningfully at a higher SDS concentration (p<0.0001). After culture of the ECM scaffold with blastema, we observed increased staining of alcian blue, periodic acid-Schiff (PAS) and toluidine blue in the scaffold and a number of other migrant cells which was caused by cell penetrationinto the scaffold. Spectrophotometry results showed an increase in glycosaminoglycans (GAGs) of the decellularized scaffolds at three weeks after culture. CONCLUSION: The present study has shown that a scaffold generated from palatal gingival tissue ECM is a suitable substrate for blastema cell migration and activity.This scaffold maypotentially be useful as a biological scaffold in tissue engineering applications.

in vitro Decellularization of Rabbit Lung Tissue.

OBJECTIVE: The bioscaffold can be used in tissue engineering and regenerative medicine. The scaffolds used in tissue engineering must have high porosity to facilitate accelerated angiogenesis for feeding cells and repelling cell waste outside the scaffold. In this experimental study, we attempted to produce lung three-dimensional scaffold and assay its effect on cell penetration and migration. MATERIALS AND METHODS: In an experimental study, rabbit lung tissue was decellularized and used as a scaffold for rabbit blastema cells. The scaffolds were studied on the 15(th) day after culturing. RESULTS: Microscopic features revealed high porosity in the lung tissue scaffold. Electron microscopic imaging also showed collagen and elastin were intact, which are important properties in scaffolds designed for tissue engineering. Migration and permeation of blastema cells into the lung tissue scaffold was also observed. CONCLUSION: Rabbit lung tissue scaffolds have high porosity. Blastema cells successfully migrated toward and permeated the scaffold inside.

Scaffolds derived from cancellous bovine bone support mesenchymal stem cells' maintenance and growth.

Since bone defects can lead to various disabilities, in recent years, many increasing attempts have been made in bone tissue engineering. In this regard, scaffolds have attracted a lot of attention as three dimensional substrates for cell attachment which improve successful tissue engineering. The aim of the present study was to provide an interconnected porous scaffold to facilitate cell infiltration. To do so, cancellous bone from bovine femur was dissected in fragments and decellularized by physicochemical methods, including snap freeze/thaw, rinsing in hot water and treatment with different solutions of sodium dodecyl sulfate (SDS). Histological analysis and 4',6-diamidino-2-phenylindole staining revealed that the best results were obtained after treatment with 2.5%, 5%, and 8% SDS for 8, 3, or 1 h respectively, which significantly removed bone cells with intact trabeculae geometry. Further characterization of decellularized scaffolds by the compression tests also revealed no significant difference between elastic modulus values of the three different SDS treatments. Moreover, studying the ratio of bone trabeculae to bone surfaces (BT/BS) as assessed by Clemex vision software 3.5 showed that treatment with 2.5% SDS for 8 h resulted in a BT/BS score in the range of native bone and therefore this treatment was used for further experiments. Histological studies and scanning electron microscopy revealed rat mesenchymal stem cells integration, adhesion, and maintenance during the 2 and 7 d of culture in vitro. In conclusion, the present results support the effective role of SDS in cancellous bovine bone decellularization and also propensity of treated samples in providing a suitable three-dimentional environment to support the maintenance and growth of mesenchymal stem cells.

Topical effects of frog "Rana ridibunda" skin secretions on wound healing and reduction of wound microbial load.

ETHNOPHARMACOLOGICAL RELEVANCE: Study of the interrelationships between human and the animals in their environment has always been a subject of interest and caused discoveries of the animal applications in medicine. From the latest century, these remedies called back in traditional medicine of Vietnam and South America and frog skin was used as a biological dressing and had good effects in healing wounds. Also, frog skin secretions have wound healing properties and reduce inflammation. In this study we applied these secretions in the form of ointment to investigate their healing activities. MATERIALS AND METHODS: Skin secretions were extracted from Rana ridibunda to evaluate their effects on wound healing in mice. Secretion used as raw extract (RE) and ultrafiltrated extract, using a membrane with cutoff 10kDa as under 10kDa (U10E), was administrated as ointment every 48h on wound site. Control group was left without any treatment and also there was other group treated with ointment (O group) alone. On 2, 4 and 6 days post injury, animals were euthanized and images were taken for wound closure evaluation. Then wound locations were removed for histological assays. Also wound microbial load was examined. Observational parameters including wound closure and wound microbiology in experimental groups compared with the control and O groups have been studied. RESULTS: The results showed U10E group has better effects than RE, O and control groups. Histological parameters, including numbers of inflammatory and fibroblast cells and amount of collagen fibers, neovascularization, as well, represented greater degree of wound healing in U10E group compared with RE, O, and control groups. CONCLUSIONS: Our results showed that frog skin secretions were significantly effective in promoting wound healing process. The U10E extract from the frog R. ridibunda possesses a potent accelerating wound healing effect that promises good potential for clinical application in wound care. Further studies will be required to characterize special molecules encompassing healing properties.