The Combined Effect of Lumenato and Ceramide in the Protection of Collagen Damage Induced by Neutrophils in Normal Human Dermal Fibroblasts

Introduction: Collagen is the primary structural protein fibroblasts produce in the skin’s extracellular matrix. Infiltration of neutrophils into the epidermis and dermis by exposure to UV causes collagen damage and contributes to photoaging. Methods: To study the combined effect of Lumenato and ceramide in preventing collagen-1 damage induced by phagocytes, we used co-cultures of normal human dermal fibroblasts (fibroblasts) and activated human neutrophils. The present study aimed to determine the protective effect of the combination of Lumenato and ceramide on fibroblast collagen-1 damage induced by neutrophils. Results: Lumenato (in the range of 6.5 - 208 μg/ml) or ceramide (in the range of 0.1 - 50 μM) inhibited the production of superoxides and MPO by TNFα-stimulated neutrophils, as well as the production of NO by LPS-stimulated macrophages in a dose-dependent manner. The combinations of Lumenato and ceramide, in low concentrations, caused synergistic prevention of fibroblasts’ collagen-1 damage induced by TNFα-activated neutrophils, detected by fluorescence immunostaining and WB analysis. MPO activity in the supernatants of the co-cultures was also synergistically inhibited. Adding Lumenato or ceramide singly or in combinations in these low concentrations to the fibroblast cultures did not affect the expression of collagen-1. The combinations of Lumenato or ceramide in these concentrations also caused a synergistic inhibition of NO production by activated macrophages. Conclusions: The results suggest that combining low concentrations of Lumenato and ceramide results in synergistic protection against fibroblasts’ collagen-1 damage induced by neutrophils, thus indicating their possible potential for enhanced skin health.

Lysine demethylase 5B transcriptionally regulates TREM1 in human cardiac fibroblasts

Background:A differential gene,triggering receptor expressed on myeloid cells 1(TREM1),was identified in blood sequencing datasets from myocardial infarction patients and healthy controls.Myocardialfibrosis following myocardial infarction significantly contributes to cardiac dysfunction.Objectives:This study aimed to unveil the intrinsic regulatory mechanism of TREM1 in myocardialfibrosis.Methods:Mimicking pathology by angiotensin II(Ang II)treatment of human cardiacfibroblasts(HCFs),the impacts of TREM1 knockdown on its proliferation,migration,and secretion of the pro-fibrotic matrix were identified.Using the Human Transcription Factor Database(HumanTFDB)website,lysine-specific demethylase 5B(KDM5B)was found to bind to the TREM1 promoter,which was further validated through luciferase reporter and chromatin immunoprecipitation(ChIP).By promoting KDM5B overexpression,its effect on the regulation of TREM1 was examined.Results:TREM1 knockdown suppressed the proliferation,migration,and secretion of the pro-fibrotic matrix in HCFs upon Ang II treatment.KDM5B bound to the TREM1 promoter and upregulated its transcriptional expression.Furthermore,KDM5B overexpression reversed the regulation of the above cellular phenotypes by TREM1 knockdown.Conclusion:This study sheds light on the positive regulation of TREM1 by KDM5B,demonstrating their role in promoting myocardialfibrosis.Thisfinding provides a theoretical foundation for understanding disease pathology and potentially advancing the development of new targeted therapies.

17β-estradiol inhibits TGF-β-induced collagen gel contraction mediated by human Tenon fibroblasts via Smads and MAPK signaling pathways

AIM:To investigate the impact of 17β-estradiol on the collagen gels contraction(CGC)and inflammation induced by transforming growth factor(TGF)-βin human Tenon fibroblasts(HTFs).METHODS:HTFs were three-dimensionally cultivated in type I collagen-generated gels with or without TGF-β(5 ng/mL),17β-estradiol(12.5 to 100μmol/L),or progesterone(12.5 to 100μmol/L).Then,the collagen gel diameter was determined to assess the contraction,and the development of stress fibers was analyzed using immunofluorescence staining.Immunoblot and gelatin zymography assays were used to analyze matrix metalloproteinases(MMPs)and tissue inhibitors of metalloproteinases(TIMPs)being released into culture supernatants.Enzyme-linked immunosorbent assay(ELISA)and reverse transcription-quantitative polymerase chain reaction(RT-PCR)were used to detect interleukin(IL)-6,monocyte chemoattractant proteins(MCP)-1,and vascular endothelial growth factor(VEGF)in HTFs at the translational and transcriptional levels.The phosphorylation levels of Sma-and Mad-related proteins(Smads),mitogen-activated protein kinases(MAPKs),and protein kinase B(AKT)were measured by immunoblotting.Statistical analysis was performed using either the Tukey-Kramer test or Student’s unpaired t-test to compare the various treatments.RESULTS:The CGC caused by TGF-βin HTFs was significantly inhibited by 17β-estradiol(25 to 100μmol/L),and a statistically significant difference was observed when comparing the normal control group with 17β-estradiol concentrations exceeding 25μmol/L(P<0.05).The suppressive impact of 17β-estradiol became evident 24h after administration and peaked at 72h(P<0.05),whereas progesterone had no impact.Moreover,17β-estradiol attenuated the formation of stress fibers,and the production of MMP-3 and MMP-1 in HTFs stimulated by TGF-β.The expression of MCP-1,IL-6,and VEGF mRNA and protein in HTFs were suppressed by 100μmol/L 17β-estradiol(P<0.01).Additionally,the phosphorylation of Smad2 Smad3,p38,and extracellular signal-regulated kinase(ERK)were downregulated(P<0.01).CONCLUSION:17β-estradiol significantly inhibits the CGC and inflammation caused by TGF-βin HTFs.This inhibition is likely related to the suppression of stress fibers,inhibition of MMPs,and attenuation of Smads and MAPK(ERK and p38)signaling.17β-estradiol may have potential clinical benefits in preventing scar development and inflammation in the conjunctiva.

Anti-aging Effects of Alu Antisense RNA on Human Fibroblast Senescence Through the MEK-ERK Pathway Mediated by KIF15

Objective:To investigate whether human short interspersed nuclear element antisense RNA(Alu antisense RNA;Alu asRNA)could delay human fibroblast senescence and explore the underlying mechanisms.Methods:We transfected Alu asRNA into senescent human fibroblasts and used cell counting kit-8(CCK-8),reactive oxygen species(ROS),and senescence-associated beta-galactosidase(SA-β-gal)staining methods to analyze the anti-aging effects of Alu asRNA on the fibroblasts.We also used an RNA-sequencing(RNA-seq)method to investigate the Alu asRNA-specific mechanisms of anti-aging.We examined the effects of KIF15 on the anti-aging role induced by Alu asRNA.We also investigated the mechanisms underlying a KIF15-induced proliferation of senescent human fibroblasts.Results:The CCK-8,ROS and SA-β-gal results showed that Alu asRNA could delay fibroblast aging.RNA-seq showed 183 differentially expressed genes(DEGs)in Alu asRNA transfected fibroblasts compared with fibroblasts transfected with the calcium phosphate transfection(CPT)reagent.The KEGG analysis showed that the cell cycle pathway was significantly enriched in the DEGs in fibroblasts transfected with Alu asRNA compared with fibroblasts transfected with the CPT reagent.Notably,Alu asRNA promoted the KIF15 expression and activated the MEK-ERK signaling pathway.Conclusion:Our results suggest that Alu asRNA could promote senescent fibroblast proliferation via activation of the KIF15-mediated MEK-ERK signaling pathway.

Effect of a Nutrient Mixture on Fanconi Anemia Fibroblast and Normal Human Dermal Fibroblast: A Comparison

Fanconi anemia (FA) is a fatal heterogeneous autosomal recessive disorder, characterized by progressive bone marrow failure, congenital defect and cancer predisposition. Cell culture from FA fibroblast (FAF) displays certain abnormalities as compared to normal human dermal fibroblast (NHDF). This prompted us to investigate the effect of a specific nutrient mixture (NM) containing ascorbic acid, lysine, proline and green tea extract, which has demonstrated a broad spectrum of pharmacological activities, on FAF compared to NHDF. We investigated the in vitro effect of NM on FAF and NHDF cell proliferation by MTT assay, MMPs secretion by zymography, morphology by H&E staining and apoptosis by green caspase assay. FAF (FA-A: PD20, FA-A: PD220) and NHDF were cultured in modified Dulbecco Eagle media. At near confluence, the cells were treated with different concentrations of NM (0, 50, 100, 250, 500 and 1000 μg/ml) in triplicate. The cells were also treated with PMA to induce MMP-9 activity. NM had no effect on FAF cell viability in both cell lines compared to control. In contrast NM exhibited 20% at 50 and 100, 50% at 250, 60% at 500 and 70% toxicity at 1000 μg/ml on NHDF cells. Zymography demonstrated MMP-2 and MMP-9 on PMA stimulation in FAF and NM inhibited the activity of both MMP-2 and MMP-9 in a dose response fashion with total block at 500 μg/ml. In contrast, NHDF exhibited only MMP-2, both active and inactive forms, and NM inhibited their activities in a dose-dependent manner with total block at 1000 μg/ml. H&E staining did not indicate any morphological changes in FAF nor induced apoptosis at higher concentrations, as seen by caspases assay. However, although no morphological changes in NHDF were noted up to NM 100 μg/ml, progressive changes in cell shrinkage, rounding and nuclear condensation, pertaining to apoptosis, were observed at higher concentrations. These changes were consistent with the results from the green caspases apoptosis assay. Our data demonstrate that NM exhibited different responses toward FAF and NHDF. This may in part be due to elevated chromosomal break, deletion and hypersensitivity to cross linking agents, a DNA repair disorder in FAF that is lacking in NHDF.

Expression of fibroblast activation protein in human pancreatic adenocarcinoma and its clinicopathological significance

AIM: To examine fibroblast activation protein (FAP) expression in pancreatic ductal adenocarcinoma (PDAC) and to analyze its relationship with the clinicopathology of PDAC. METHODS: FAP expression was examined in 134 PDAC specimens by immunohistochemistry, and in four pancreatic cancer cell lines (SW1990, Miapaca-2, AsPC-1 and BxPC-3) by Western blotting assay. We also analyzed the association between FAP expression in PDAC cells and the clinicopathology of PDAC patients. RESULTS: The results showed that the FAP was expressed in both stromal fibroblast cells (98/134, 73.1%) and carcinoma cells (102/134, 76.1%). All 4 pancreatic cancer cell lines expressed FAP protein at different levels. Protein bands corresponding to the proteolytically active 170-kDa seprase dimer and its88-kDa seprase subunit were identif ied. Higher FAP expression in carcinoma cells was associated with tumor size (P < 0.001), fi brotic focus (P = 0.003), perineural invasion (P = 0.009) and worse clinical outcome (P = 0.0085). CONCLUSION: FAP is highly expressed in carcinoma cells and f ibroblasts in PDAC tissues, and its expression is associated with desmoplasia and worse prognosis.

Protective effects of non-mitogenic human acidic fibroblast growth factor on hydrogen peroxide-induced damage to cardiomyocytes in vitro

AIM： To study the protective effect of non-mitogenic human acidic fibroblast growth factor （FGF） on cardiac oxidative injury in vivo. METHODS： Ventricular cardiomyocytes were isolated from 1- to 3-d-old neonatal SD mice and cultured in Dulbecco＇s minimum essential medium supplemented with 15% fetal bovine serum under an atmosphere of 50 mL/L CO2-95% air at 37℃, as well as assessed by immunooltochemical assay. We constructed the cardiomyoolte injury model by exposure to a certain concentration of H2O2. Cellular viability, superoxide dismutase （SOD） activity, leakage of maleic dialdehyde and anti-apoptosis effect were included to evaluate the cardiac protective effect of non-mitogenic human acidic FGF. RESULTS： Over 50% of the cardiomyocytes beat spontaneously on the 2nd d of culture and synchronously beat after being cultured for 3 d. Forty-eight hours after plating was completed, the purity of such cultures was 95% myocytes, assessed by an immunocytochemical assay. Cellular viability dramatically decreased with the increasing of the concentration of H2O2. Non-mitogenic human acidic FGF showed significant resistance to thetoxic effect of H2O2, significantly increased the cellular viability as well as the activity of SOD, and dramatically decreased the leakage of maleic dialdehyde as well as the cellular apoptosis rate. CONCLUSION： Hydrogen peroxide shows strong cytotoxicity to the cultured cardiac myocytes, and non-mitogenic human acidic FGF shows strong cardio-protective effect when exposed to a certain concentration of H2O2.

Basic Fibroblast Growth Factor and Fibroblast Growth Factor Receptor-1 in Human Meningiomas

The expression of basic fibroblast growth factor (bFGF) and fibroblast growth factor receptor-1 (FGFR-1) in human meningiomas and the relationships between their expression and the tumors' histological features and angiogenesis were investigated by means of immunohistochemical technique. The expression of bFGF and FGFR-1 was detected by antibody of bFGF or FGFR-1. The tumors' angiogenesis was evaluated by microvascular density (MVD) and, which was observed by use of CD34-antibody immunohistochemically. The results showed that there were varied degrees of the expression of bFGF and FGFR-1 proteins in meningiomas. The expression was correlated with the tumors' histological characters and angiogenesis. It was concluded that bFGF and FGFR-1 might play important roles in meningiomas' angiogenesis and proliferation. The expression positive rate of bFGF and FGFR-1 may provide an indication of evaluating the histological and malignant degree of the tumor.

Ceruloplasmin or Fibronectin Synergism with Quartz Dust on Stimulating Collagen Gene Transcription in Human 2BS Fibroblast

Human α1(Ⅰ), α2(Ⅰ) and α1(Ⅲ) cDNA probes and RNA dot hybridization were employed to quantitate collagen mRNA changes after adding silica dust into the media of human 2BS fibroblasts. At all dosages used (100, 200, 500 and 1000μg), the α1(Ⅰ), α2(Ⅰ)and α1(Ⅲ) mRNA levels increased one day after dusting. At the same dosage of silica (100μg), α1(Ⅲ) mRNA increased earlier than type Ⅰ collagen mRNA did. The type Ⅰ and type Ⅲ collagen mRNA contents in the experimental groups were higher than those in control on days 3, 5, 7 and 9. The effect of ceruloplasmin (Cp) and fibronectin (Fn) on collagen mRNA synthesis was also studied, after adding silica dust, Cp or Fn into the media of human 2BS fibroblast. The results showed that Cp and Fn have stimulating effect on collagen mRNA production. When both Cp and silica dust were added into cell culture media, the collagen mRNA level was increased more than those of adding either Cp or silica dust alone. Similar situations were found for Fn. Cp (or Fn) synergism with silica dust on stimulating transcription of human collagen gene was suggested

EFFECTS OF TRANSFORMING GROWTH FACTOR β AND RECOMBINANT HUMAN BONE MORPHOGENETIC PROTEIN 2 ON HUMAN PERIODONTAL LIGAMENT FIBROBLASTS

Objective To evaluate the effects of transforming growth factor β(TGF-β) and recombinant human bone morphogenetic protein 2 (rhBMP2) on human periodontal ligament fibroblasts (HPDLFs). Methods HPDLFs were done primary culture to detect the distinct concentrations of TGF-P and rhBMF2 on its proliferation, alkaline phosphatase (ALP) activity, osteocalcin (OC) synthesis and formation of the minerali-zed nodules, respectively. Results TGF-β (5～100ng/ml) significantly stimulated the proliferation of HPDLFs. The ALP activity of HPDLFs was evaluated evidently by 5ng/ml TGF-β. TGF-β( 0. 5 ～ 100ng/ml) had no effects on OC synthesis and formation of the mineralized nodules of HPDLFs. rhBMP2 (0. 25～2mg/ ml) had no remarkable effect on the proliferation of HPDLFs. The ALP activity, OC synthesis and forma-tion of the mineralized nodules of HPDLFs were significantly stimulated by 0. 5～ 2mg /ml rhBMP2. Conclusion The effects of TGF-β and rhBMP2 on HPDLFs are dose-dependent. TGF-P can stimulate HPDLFs to express the early marker of osteoblastic phenotype, and it lacks the ability to promote maturation of the osteogenic phenotype. rhBMP2 can not only stimulate the expression but also promote the maturation of osteoblas-tic phenotype of HPDLFs.

Experimental study on effect of endothelin in the proliferation and collagen synthesis of human scar-derived fibroblasts

Objective To investigate the role of endothelin(ET) in the proliferation and collagen synthesis of human scar-derived fibroblasts and the moduktion of its antagonists such as nitric oxide(NO), tetrandrine ( Tet). Methods With the cultured fibroblasts from the scarring tissue, the cell pdiferation was determined by[3H]-TdR incorporation, while the collagen synthesis was evaluated by[3H]-proline incorporation. Results The ET-1 was significantly increasing the proliferation and collagen synthesis of human scar-derived fibroblasts. The values of [3H]-TdR absorption in the 2.5 ng/ml,25 ng/ml and 100 ng/ml of ET-1 groups were 1.8 times,4 times and 4.9 times more than in the control group, respectively(P【0. 01),while the values of the [3H]-proline incorporation were 1.1 times,3.1 times and 3.8 times respectively(P【0.01). The fibroblasts, treated with 50 μg/ml of S-nitroso-N-acetyl penicillamine(SNAP), were no detectable effect on the basal level of DNA synthesis,but produced decreasing effect on the

Inhibitory Effect of Curcumin on Proliferation of Human Pterygium Fibroblasts

In order to investigate the effect of curcumin on proliferation and apoptosis of human pterygium fibroblasts (HPF) in culture and search for a new method to prevent the recurrence after pterygium surgery, HPF was incubated with 0-160 μmol/L curcumin for 24-96 h. The MTT method was used to assay the biologic activities of curcumin at different time points and different doses. The expression of proliferating cell nuclear antigen (PCNA) in each group was detected by immunohistochemistry. The cell cycle distribution was detected by flow cytometry (FCM). Admini- stration of 20-80 μmol/L curcumin for 24-72 h could significantly inhibit HPF proliferation in a dose- and time-dependent manner (P<0.05). After treatment with curcumin at different concentrations of 20, 40, 80 and 160 μmol/L for 24 h, FCM revealed there was a significant sub-G1 peak at each concentration. The number of HPF in G0/G1 phase was increased, while in S phase, it was decreased (P<0.05). At the concentration of 20-80 μmol/L, curcumin, in a dose-dependent manner (P<0.05), could inhibit the expression of PCNA in HPF. It was suggesterd that curcumin could significantly in- hibit the proliferation of HPF, make HPF arrest in G0/G1 phase and induce the apoptosis of HPF in a dose- and time-dependent manner.

Distribution of bone morphogenetic protein receptors in human scleral fibroblasts cultured in vitro and human sclera

AIM: To investigate the distribution of bone morphogenetic protein receptors (BMPRs) in human scleral fibroblsasts (HSFs) and in human sclera. METHODS: Primary HSFs were cultured in vitro. The mRNA levels of BMP-2 and BMPRs in HSFs were assayed by reverse transcription-polymerase chain reaction (RT-PCR). The protein distributions of BMP-2 and BMPRs in HSFs were further detected by immunocytofluorescence and western blot. Their protein expression was also detected in frozen human posterior scleral sections by immunohistofluorescence. RESULTS: BMP-2 and BMPRs were expressed in both HSFs and human sclera not only at mRNA level but also at protein level. The expressions of BMPRIA and BMPRII were higher than that of BMPRIB in the cytoplasm and cell membrane of HSFs in vitra Western blot further verified the results of immunocytofluorescence. In human sclera, BMP2, BMPR IB and BMPR II were found to be expressed in the cytomatrix of HSF, and weak signal was detected about BMPRIA. CONCLUSION: BMP-2 and all three subtypes of BMPRs were found in HSFs and may play a role in scleral remodeling.

Over-expression of fibroblast growth factor receptor 3 in human hepatocellular carcinoma

AIM: To describe the significant over-expression of fibroblast growth factor receptor 3 (FGFR3), which is a signal transduction and cell proliferation related gene in hepatocellular carcinoma (HCC).METHODS: Following DNA microarray, Northern blot and quantitative real-time PCR were employed to confirm FGFR3 expression difference in HCC tissues and surrounding non-neoplastic liver tissue. FGFR3 expression levels were further determined by immunohistochemical study in 43 cases of HCC.RESULTS: Northern blot results showed the significant over-expression of FGFR3 in HCC tissues, which was consistent with that from DNA microarray. Quantitative real-time PCR demonstrated that the mean ratio of FGFR3 mRNA to glyceraldehyde-3-phosphate dehydrogenase (GADPH) mRNA in HCC tissue was 0.250, whereas the ratio in non-neoplastic liver tissue was 0.014. Statistical analyses of 43 cases of HCC revealed that HCC scored higher than the matched non-neoplastic liver tissues.Examination of clinicopathological features revealed a strong correlation of over-expression of FGFR3 with poor tumor differentiation and high nuclear grade.CONCLUSION: Over-expression of FGFR3 may play an important role in liver carcinogenesis. FGFR3 may be an ideal candidate as a molecular marker in the diagnosis of HCC and a potential therapeutic target.

Noggin versus basic fibroblast growth factor on the differentiation of human embryonic stem cells

The difference between Noggin and basic fibroblast growth factor for the neural precursor differen- tiation from human embryonic stem cells has not been studied. In this study, 100 tJg/L Noggin or 20 IJg/L basic fibroblast growth factor in serum-free neural induction medium was used to differen- tiate human embryonic stem cells H14 into neural precursors using monolayer differentiation. Two weeks after induction, significantly higher numbers of neural rosettes formed in the Noggin-induced group than the basic fibroblast growth factor-induced group, as detected by phase contrast micro- scope. Immunofluorescence staining revealed expression levels of Nestin, [3-111 Tubulin and Sox-1 were higher in the induced cells and reverse-transcription PCR showed induced cells expressed Nestin, Sox-1 and Neurofilament mRNA. Protein and mRNA expression in the Noggin-induced group was increased compared with the basic fibroblast growth factor-induced group. Noggin has a greater effect than basic fibroblast growth factor on the induction of human embryonic stem cell differentiation into neural precursors by monolayer differentiation, as Noggin accelerates and in- creases the differentiation of neural precursors.

An In-depth Interpretation of the Universal Declaration of Human Rights With the Common Values of Humanity As the Research Paradigm

Interpreting the Universal Declaration of Human Rights from political,juridical and philosophical perspectives is es-sential for promoting the guiding principles of the Declaration,build-ing consensus on human rights,and advancing human rights practice in the new historical context.To conduct an academic,systematic in-terpretation of the Declaration that conforms to the trends of the times and answers the fundamental questions of the world,it is necessary to find a new research paradigm.The common values of humanity,namely peace,development,equity,justice,democracy and freedom,put forward by Xi Jinping,general secretary of the Communist Party of China(CPC)Central Committee,provide the most explanatory and penetrating scientific paradigm for reaching the issue.This paper an-alyzes and reflects on the views,value foundation and principled(con-tractual)consensus of human rights in the Declaration,and narrates and foresees the far-reaching significance of the three global initia-tives(namely,the Global Development Initiative,the Global Security Initiative,and the Global Civilization Initiative)with the common val-ues of humanity as the soul in advancing the modernization of global human rights governance and building a new form of human rights civilization.

Effects of 13 T Static Magnetic Fields (SMF) in the Cell Cycle Distribution and Cell Viability in Immortalized Hamster Cells and Human Primary Fibroblasts Cells

Magnetic resonance image （MRI） systems with a much higher magnetic flux density were developed and applied for potential use in medical diagnostic. Recently, much attention has been paid to the biological effects of static, strong magnetic fields （SMF）. With the 13 T SMF facility in the Institute of Plasma Physics, Chinese Academy of Sciences, the present study focused on the cellular effects of the SMF with 13 T on the cell viability and the cell cycle distribution in immortalized hamster cells, such as human-hamster hybrid （AL） cells, Chinese hamster ovary （CHO） cells, DNA double-strand break repair deficient mutant （XRS-5） cells, and human primary skin fibroblasts （AG1522） cells. It was found that the exposure of 13 T SMF had less effect on the colony formation in either nonsynchronized or synchronized AL cells. Moreover, as compared to non-exposed groups, there were slight differences in the cell cycle distribution no matter in either synchronized or nonsynchronized immortalized hamster ceils after exposure to 13 T SMF. However, it should be noted that the percentage of exposed AG1522 cells at G0/G1 phase was decreased by 10% as compared to the controls. Our data indicated that although 13 T SMF had minimal effects in immortalized hamster cells, the cell cycle distribution was slightly modified by SMF in human primary fibroblasts.

ETM study of electroporation influence on cell morphology in human malignant melanoma and human primary gingival fibroblast cells

Objective:To estimate electroporation(EP) influence on malignant and normal cells.Methods: Two cell lines including human malignant melanoma(Me-43) and normal human gingival fibroblast(HCFs) were used.EP parameters were the following:230,1000,1 730,2 300 V/cm;30 μ s by 3 impulses for every case.The viability of cells after EP was estimated by MTT assay. The ullrastructural analysis was observed by transmission electron microscope(Zeiss EM 900). Results:In the current study we observed the intracellular effect following EP on Me-43 and HGF cells.At the conditions applied,we did not observe any significant damage of mitochondrial activity in both cell lines treated by EP.Conversely,we showed that EP in some conditions can stimulate cells to proliferation.Some changes induced by EP were only visible in electron microscopy.In fibroblast cells we observed significant changes in lower parameters of EP(230 and 1 000 V/cm).After applying higher electric field intensities(2 300 V/cm) we detected many vacuoles,myelin-like bodies and swallowed endoplasmic reticulum.In melanoma cells such strong pathological modifications after EP were not observed,in comparison with control cells. The ultrastructure of both treated cell lines was changed according to the applied parameters of EP.Conclusions:We can claim that EP conditions are cell line dependent.In terms of the intracellular morphology,human fibroblasts are more sensitive to electric field as compared with melanoma cells.Optimal conditions should be determined for each cell line.Summarizing our study,we can conclude that EP is not an invasive method for human normal and malignant cells. This technique can be safely applied in chemotherapy for delivering drugs into tumor cells.

ZnO,TiO_2,SiO_2,and Al_2O_3 Nanoparticles-induced Toxic Effects on Human Fetal Lung Fibroblasts

Objective This study aims to investigate and compare the toxic effects of four types of metal oxide （ZnO, TiO2, SiO2, and Al2O3） nanoparticles with similar primary size （-20 nm） on human fetal lung fibroblasts （HFL1） in vitro.Methods The HFL1 cells were exposed to the nanoparticles, and toxic effects were analyzed by using MTT assay, cellular morphology observation and Hoechst 33 258 staining.Results The results show that the four types of metal oxide nanoparticles lead to cellular mitochondrial dysfunction, morphological modifications and apoptosis at the concentration range of 0.25-1.50 mg/mL and the toxic effects are obviously displayed in dose-dependent manner. ZnO is the most toxic nanomaterials followed by TiO2, SiO2, and Al2O3 nanoparticles in a descending order.Conclusion The results highlight the differential cytotoxicity associated with exposure to ZnO, TiO2, SiO2, and Al2O3 nanoparticles, and suggest an extreme attention to safety utilization of these nanomaterials.

Chicken collagen hydrolysates differentially mediate anti-inflammatory activity and type I collagen synthesis on human dermal fibroblasts

Collagen is a major extracellular matrix protein.Given the potential anti-inflammatory and antioxidant profiles of these bioactive compounds,there has been increasing interest in using collagen derived peptides and peptide-rich collagen hydrolysates for skin health,due to their immunomodulatory,antioxidant and proliferative effects on dermal fibroblasts.However,all hydrolysates are not equally effective in exerting the beneficial effects;hence,further research is needed to determine the factors that improve the therapeutic applicability of such preparations.We used different enzymatic conditions to generate a number of different collagen hydrolysates with distinct peptide profiles.We found that the use of two rather than one enzyme for hydrolysis generates a greater abundance of low molecular weight peptides with consequent improvement in bioactive properties.Testing these hydrolysates on human dermal fibroblasts showed distinct actions on inflammatory changes,oxidative stress,type I collagen synthesis and cellular proliferation.Our findings suggest that different enzymatic conditions affect the peptide profile of hydrolysates and differentially regulate their biological activities and potential protective responses on dermal fibroblasts.