Selective Excitation of the 1Lα State of Tryptophan in Collagen-like Peptides Can Reveal the Formation of a Heterotrimer.

Fluorescence emission from tryptophan residues has been often used to probe the protein structure due to its transition dipole moment sensitivity to the local environment. We report the fluorescence study of a collagen-like peptide heterotrimer modified with the tryptophan in the X position (X-Y-Gly) n that shows the diminished fluorescence in a homotrimer versus a heterotrimer when the 1Lα state is selectively excited. This behavior is only observed in folded peptides, below the helix-to-coil transition temperature, and can be explained by long-range interactions between the tryptophans located on different strands within the triple helix, not by the change in the local environment. Our results suggest that tryptophan homotransfer is possible at distances much longer than the R 0 (0.5-0.7 nm) previously estimated. These observations imply that the energy transfer between the 1Lα states of proximal tryptophans can be facilitated by constraining their rotation by the helix and, thus, can be employed as a reporter of heterotrimer formation in biosensors.

Hybrid peptides as platform for synchronized combination therapy.

Combination therapy, where two or more therapeutic agents are combined to target different cellular pathways, is an effective tool in cancer treatment but often difficult to execute. Here we present the collagen peptide-based platform that allows for synchronous and colocalized cellular delivery of three different agents. The peptide is a hybrid between collagen and cell penetrating peptide (CPP) that assembles into a heterotrimer helix and forms fully organic, high aspect ratio nanoparticles. The validity of the approach was tested with three chemically different agents (Paclitaxel, Doxorubicin, and 5-Fluorouracil; a combination used in clinical treatment of (ER)-positive and (PR)-positive breast cancer) conjugated to N-terminus of the peptide. The design of this peptide-based drug delivery system provides several advantages: it avoids drug loading problems; removes the need for orthogonal synthesis; and allows for colocalized delivery of up to three drugs (which leads to the same biodistribution for each drug). In addition, hybrid collagen/CPP peptides are known to enhance cellular uptake and improve solubility of drugs. The synergistic effect, in terms of enhanced efficacy, of the Paclitaxel-Doxorubicin-5-Fluorouracil combination was also calculated. We envision self-assembling peptides as a platform for drug codelivery that can be expanded into a library of personalized combinations that may also include other functionalities like targeting or imaging.

Dietary Protein Source Matters for Changes in Inflammation Measured by Urinary C-Reactive Protein in Rural Polish Women.

Objectives: Multiple macronutrients have been shown to affect systemic inflammation, a well-known predictor of chronic disease. Less often, varying sources of these macronutrients are examined. Different subsistence environments lead to varying access to protein sources which, combined with physical activity patterns, may lead to different relationships than among more typically studied sedentary, industrialized populations. This study hypothesizes an association between dietary protein intake and urinary C-Reactive Protein (CRP) concentration in women from a rural, agrarian Polish community. Materials and Methods: We assessed protein intake and their sources for 80 nonsmoking, premenopausal Polish women who were not pregnant, nursing, or on hormonal birth control during the study or within the previous six months. Each participant completed multiple 24-hour dietary recalls during one menstrual cycle. Participants collected morning void urinary samples daily over one menstrual cycle for urinary CRP analysis. We analyzed relationships between plant and animal protein intake and CRP over the menstrual cycle by multiple linear regression. Results: Plant protein in cereal foods was significantly positively associated with cycle-average urinary CRP concentrations (p<0.05) after controlling for body fat percent, total energy intake, and dietary fiber. Foods containing animal protein were not significantly associated with CRP. Discussion: Contents of this population's main plant and animal protein sources differ from those of more commonly studied industrialized populations. Within the context of a population's typical diet, more emphasis may need to be placed on particular source of protein consumed, beyond plant versus animal, in order to understand relationships with CRP.

Position statement of the Polish Academy of Sciences Committee Human Nutrition Science on the principles for the nutrition of children aged 1-3 years.

The Position Statement on the principles of nutrition for children aged 1-3 years emphasizes that proper nutrition of children at this age determines their optimal psychometric development and has beneficial effects on the process nutritional programming, which reduces the risk of diet-related diseases in adulthood. Continued breastfeeding in the post-infancy period, together with the proper introduction of complementary foods, supplies all the nutritional needs of the child. A varied selection of food products is important to balance out the diet of a child in the context of energy and nutrient needs. Attention should be paid to products not recommended for frequent consumption, due to the possibility of the early development of improper eating habits that can lead to undesirable health consequences. Due to the potential risk of deficiency, adequate intake of iron, iodine, calcium and vitamin D, as well as of n-3 PUFAs (which is often insufficient) should be provided. Adequate dietary energy and protein intake protects children against protein-energy undernutrition and is crucial for their proper growth and development. An important element in the assessment of the development of children involves monitoring their nutritional status and physical development by systematically measuring their body weight and length/height and analyzing their weight gain. It is necessary to diagnose the causes of being underweight/overweight in children. Physical activity (such as outdoor walks, plays, and games) and healthy sleep hygiene are recommended. Physical activity, an adequate number of hours of sleep, and the quality of sleep in early childhood may improve immunity, reduce the risk of excessive weight gain, and consequently reduce the risk of obesity later in life. Other issues discussed include the functioning of the digestive system as one of the determinants of the nutrition of young children, basics of proper nutrition, risk of nutrient deficiencies and development of proper eating habits in early childhood.

Serum Total SOD Activity and SOD1/2 Concentrations in Predicting All-Cause Mortality in Lung Cancer Patients.

Redox status disturbances are known during carcinogenesis and may have influence on patients' survival. However, the prediction of mortality in lung cancer patients based on serum total SOD activity, and concentrations of its isoforms, has not been studied to date. This prospective cohort study has following aims: (1) to evaluate the disturbances in serum SOD activity and SOD1/2 concentrations; (2) to assess the implications of these alterations with regard to biochemical variables and clinical data, and (3) to investigate the association between serum SOD activity, SOD1/2 concentrations, and all-cause mortality in lung cancer patients. Serum total SOD activity and SOD1, SOD2, albumin, CRP, and ceruloplasmin concentrations were determined in lung cancer patients (n = 190) and control subjects (n = 52). Additionally, patients were characterized in terms of biochemical, clinical, and sociodemographic data. Multiple Cox regression models were used to estimate the association between all-cause death and SOD-related parameters. All-cause mortality in lung cancer was positively associated with serum SOD1 and SOD2 concentrations. Clinical stage III and IV disease was the strongest predictor. The utility of the evaluated parameters in predicting overall survival was demonstrated only for SOD1. Serum SOD1 and SOD2 concentrations were shown to positively affect all-cause mortality in lung cancer patients, but SOD1 seems to be a better predictor than SOD2.

Frequency of legume consumption related to sociodemographic factors, health status and health-related variables among surveyed adults from Poland.

OBJECTIVE: The aims of the study were: (i) to estimate the frequency of legume consumption in a study sample from Poland; (ii) to assess relationships between frequency of legume consumption and sociodemographic factors, health status and health-related variables and (iii) to describe the sociodemographic and health-related profiles of 'frequent', 'rare' and 'never legume consumers'. DESIGN: A cross-sectional study with a self-administered questionnaire was conducted. χ2 test compared the distribution of sociodemographic, health status and health-related variables in accordance with frequency of legume consumption. Multiple correspondence analysis (MCA) revealed the sociodemographic and health-related profiles of 'frequent', 'rare' and 'never legume consumers'. SETTING: Adult individuals from Poland. PARTICIPANTS: The present study included 1135 adults aged 18-80 years. RESULTS: Only 22·8 % of all participants declared frequent legume consumption. The prevalence of 'rare' and 'never consumers' was 72·1 and 5·1 %, respectively. The MCA highlighted strong correspondence of sociodemographic factors, health status and health-related variables with the frequency of legume consumption. CONCLUSIONS: Low frequency of legume consumption in a study sample of adults from Poland was strongly associated with male sex, low educational level, physical work, living in villages and small cities, and living with more than one co-resident; low legume consumption was also associated with sedentary lifestyle, no disease prevalence and no dietary supplementation.

Serum and Whole Blood Cu and Zn Status in Predicting Mortality in Lung Cancer Patients.

Alterations in circulating Cu and Zn are negative predictors of survival in neoplastic patients and are known during lung cancer. However, no data on predicting mortality of lung cancer patients based on the level of these elements in the blood have been presented to date. The aims of this prospective cohort study were as follows: (i) To evaluate the disturbances in serum and whole blood Cu and Zn, (ii) to assess the relationships between serum and whole blood Cu and Zn status and clinical, sociodemographic, and nutritional data, and (iii) to investigate the association of Cu and Zn status with all-cause mortality in lung cancer. Naïve-treatment lung cancer patients (n = 167) were characterized in terms of sociodemographic, clinical, and anthropometric data and dietary intake and compared with sex-matched control subjects (n = 48). Whole blood and serum Cu and Zn status was determined by atomic absorption spectrometry. Cox proportional hazards models adjusted for multiple confounders/mediators were used to estimate the association between all-cause death and Cu and Zn status. Sex, cardiovascular disease, chronic obstructive pulmonary disease, clinical stage, and hemoglobin, platelet, and glucose concentrations significantly differentiated Cu and Zn status. All-cause mortality in lung cancer patients was positively associated with serum Cu levels, Cu:Zn ratio, and whole blood Zn levels. However, an advanced clinical stage of disease was the strongest predictor of all-cause mortality. Circulatory status of Cu and Zn might be included in routine clinical characteristics of patients with lung cancer patients as additional prognostic variables, but only after further more detail studies.

Tuning Crystal Structures of Iron-Based Metal-Organic Frameworks for Drug Delivery Applications.

Iron-based metal-organic frameworks (Fe-MOFs) have emerged as promising candidates for drug delivery applications due to their low toxicity, structural flexibility, and safe biodegradation in a physiological environment. Here, we studied two types of Fe-MOFs: MIL-53 and MIL-88B, for in vitro drug loading and releasing of ibuprofen as a model drug. Both Fe-MOFs are based on the same iron clusters and organic ligands but form different crystal structures as a result of two different nucleation pathways. The MIL-53 structure demonstrates one-dimensional channels, while MIL-88B exhibits a three-dimensional cage structure. Our studies show that MIL-53 adsorbs more ibuprofen (37.0 wt %) compared to MIL-88B (19.5 wt %). A controlled drug release was observed in both materials with a slower elution pattern in the case of the ibuprofen-encapsulated MIL-88B. This indicates that a complex cage structure of MIL-88 is beneficial to control the rate of drug release. A linear correlation was found between cumulative drug release and the degree of material degradation, suggesting the biodegradation of Fe-MILs as the main drug elution mechanism. The cytotoxicity of MIL-88B was evaluated in vitro with NIH-3T3 Swiss mouse fibroblasts, and it shows that MIL-88B has no adverse effects on cell viability up to 0.1 mg/mL. This low toxicity was attributed to the morphology of MIL-88B nanocrystals. The very low toxicity and controlled drug release behavior of Fe-MIL-88B suggest that better materials for drug-delivery applications can be created by controlling not only the composition but also the crystal structure and nanoparticle morphology of the material.

Declining ages at menarche in an agrarian rural region of Poland.

OBJECTIVE: Age at menarche in Poland has varied with political and socioeconomic changes. An increase in age at menarche corresponded to a period of economic crisis and food rationing between 1976 and 1989. Experiencing food shortages in utero or during childhood development can affect menarcheal timing, but this national effect may be buffered in local agrarian regions growing their own food. Here we examine patterns of age at menarche over time in the rural, agrarian Beskid Wyspowy region of southern Poland. METHODS: This study examined menarcheal timing using data collected from Polish women (n = 1326) recruited at the Mogielica Human Ecology Study Site between 2003 and 2018. Simple linear regressions were used to assess changing ages at menarche over time. Comparisons between ages at menarche for women born before and after the fall of communism in 1989 were assessed via one-way analysis of variance. RESULTS: Age at menarche has declined over time in the Beskid Wyspowy region of southern Poland from 1920 to 2000 (R2 = .08, P < .0001). There was not a statistically significant increase or decrease in age at menarche for women born and growing up during the period of food rationing. CONCLUSIONS: The declining age at menarche is likely reflective of a transitioning environment, suggesting that major socioeconomic changes affect life history traits like pubertal timing. Living in agricultural regions may have helped buffer the increasing ages at menarche seen in other areas of Poland during times of food rationing.

Higher Order Architecture of Designer Peptides Forms Bioinspired 10 nm siRNA Delivery System.

The higher-order architecture observed in biological systems, like viruses, is very effective in nucleic acid transport. The replications of this system has been attempted with both synthetic and naturally occurring polymers with mixed results. Here we describe a peptide/siRNA quaternary complex that functions as an siRNA delivery system. The rational design of a peptide assembly is inspired by the viral capsids, but not derived from them. We selected the collagen peptide (COL) to provide the structural stability and the folding framework, and hybridize it with the cell penetrating peptide (CPP) that allows for effective penetration of biological barriers. The peptide/siRNA quaternary complex forms stoichiometric, 10 nm nanoparticles, that show fast cellular uptake (<30 min), effective siRNA release, and gene silencing. The complex provides capsid-like protection for siRNA against nucleases without being immunostimulatory, or cytotoxic. Our data suggests that delivery vehicles based on synthetic quaternary structures that exhibit higher-order architecture may be effective in improving delivery and release of nucleic acid cargo.

Dietary Silicon and Its Impact on Plasma Silicon Levels in the Polish Population.

Silicon in nutritional amounts provides benefits for bone health and cognitive function. The relationship between silicon intake from a common daily diet and silicon blood level has been scarcely elucidated, so far. The aim of this study was to analyze the associations between plasma silicon levels and the total and bioavailable silicon intake-along with the contribution of silicon made by food groups-in a healthy adult Polish population. Si intake was evaluated in 185 healthy adults (94 females and 91 males, aged 20-70) using a 3-day dietary recall and a database on the silicon content in foods, which was based on both previously published data and our own research. Fasting plasma silicon levels were measured in 126 consenting subjects, using graphite furnace atomic absorption spectrometry. The silicon intake in the Polish population differed significantly according to sex, amounting to 24.0 mg/day in women and 27.7 mg/day in men. The median plasma silicon level was 152.3 µg/L having no gender dependency but with a negative correlation with age. Significant correlations were found between plasma silicon level and total and bioavailable silicon intake, as well as water intake in the diet (r = 0.18, p = 0.044; r = 0.23, p = 0.011; r = 0.28, p = 0.002, respectively). Silicon intakes from non-alcoholic beverages, cereal foods, and carotene-rich vegetables were also positively associated with plasma silicon levels. These results may help establish dietary silicon recommendations and formulate practical advice on dietary choices to ensure an appropriate supply of silicon. The outcome of this study, however, needs to be confirmed by large-scale epidemiological investigations.

Oxidative stress in lung cancer patients is associated with altered serum markers of lipid metabolism.

In lung cancer (LC), alterations in redox balance are extensively observed and are a consequence of disease as well as co-occurrent with smoking. We previously demonstrated that metabolic disturbances such as trace element status and carbohydrate metabolism alterations are linked with redox status. The aim of this study was to evaluate relationships between the serum parameters of lipid metabolism and redox balance in LC patients. Serum parameters of lipid metabolism, i.e. total cholesterol (T-C), HDL cholesterol (HDL-C), LDL cholesterol (LDL-C), triglycerides (TG), T-C:HDL-C ratio, non-HDL-C, apolipoprotein A1 (Apo-A1), apolipoprotein B (Apo-B) and Apo-B:Apo-A1 ratio, as well as systemic redox status, i.e. total antioxidant status (TAS), total oxidant status (TOS), oxidative stress index (OSI), vitamin E (VE), vitamin C (VC), malonyldialdehyde (MDA), conjugated dienes (CD), and 4-hydroxynonenal (4-HNE) were determined in 92 LC patients and 82 control subjects (CS). LC women had significantly lower T-C and LDL-C, and higher TG, while HDL-C, Apo-A1 and Apo-B were significantly decreased in LC patients regardless of sex, when compared to CS. LC men had alterations in the systemic total redox balance such as lower TAS and higher OSI than CS men. LC women had lower VC, but VE was decreased in LC patients, regardless of sex. We observed higher lipid peroxidation in LC patients expressed via higher 4-HNE and CD. Systemic redox disturbances were associated with serum lipid alterations: TOS and OSI were positively correlated with T-C:HDL-C ratio and Apo-B:Apo-A1 ratio and negatively with HDL-C. The parameters of lipid peroxidation CD and MDA were significantly associated with variables reflecting lipid disturbances. The observed correlations were strengthened by general overweight/obesity, abdominal obesity, hypertriglyceridemia and non-smoking status. In conclusion, parameters related to lipid alterations are associated with oxidative stress in LC patients. The largest contribution from lipid parameters was revealed for T-C:HDL-C ratio, HDL-C and Apo-B:Apo-A1 ratio, while the largest contribution from redox status was revealed for OSI and VE. Overweight, obesity, hypertriglyceridemia and non-smoking status intensified these relationships.

Silicon intake and plasma level and their relationships with systemic redox and inflammatory markers in rheumatoid arthritis patients.

BACKGROUND: The nutritional significance of silicon for the human body is highlighted by a continually growing body of evidence. In conditions of excessive reactive oxygen species and upregulated immune response, silicon has been observed to provide benefits, but its role in redox and inflammatory status has not yet been examined in rheumatoid arthritis (RA). OBJECTIVES: The aim of this study was to assess the relationship of silicon intake and plasma level to systemic indices of redox status and inflammation in patients with RA. MATERIAL AND METHODS: Silicon intake and plasma levels were measured in 115 RA subjects and 129 control subjects. Serum antioxidant and oxidant levels, antioxidant enzyme activity, and albumin, uric acid, TBARS, hs-CRP, and IL-6 levels were measured and compared to the intake and plasma levels of silicon. RESULTS: Silicon intake and plasma silicon levels were higher in RA subjects than in the controls. In the RA group, a generally favorable correlation to redox and inflammatory markers was found for silicon in diet and in plasma; however, albumin level, smoking status, and gender interfered with these results. In the control subjects, a significant relationship was observed only between plasma silicon and non-enzymatic markers of redox status. CONCLUSIONS: There are suggestions of silicon's involvement in managing redox and inflammatory status in RA, though further studies are warranted.

The relationships between glycemic index and glycemic load of diets and nutritional status and antioxidant/oxidant status in the serum of patients with lung cancer.

BACKGROUND: A low glycemic index (GI) and glycemic load (GL) of diets as well as proper nutritional status may partially slow down depletion in antioxidant capacity, and may therefore have an impact on antioxidant/ oxidant status in lung cancer patients. However, no studies concerning these associations had previously been conducted. OBJECTIVES: The aim of this study was to investigate the association between GI or GL and nutritional status and antioxidant/oxidant status in lung cancer patients. MATERIAL AND METHODS: The study was conducted among 180 lung cancer patients (82 women and 98 men) and 171 control subjects (78 women and 93 men). Exclusion criteria for the control subjects included cancers, pro-inflammatory conditions, brain diseases, and psychiatric disorders. All participants were evaluated in terms of their systemic antioxidant/oxidant status, nutritional status (anthropometric parameters), dietary GI and GL and parameters related to circulating glucose: fasting glucose, insulin level and homeostasis model assessment for insulin resistance (HOMA-IR). RESULTS: In women who were lung cancer-positive, associations were observed between total antioxidant status (TAS) and parameters of nutritional status, and between oxidative stress index (OSI) and fasting glucose. In men with lung cancer, we found a positive correlation between total oxidant status (TOS) and GI. In the control group of women, TAS positively correlated with anthropometric parameters, but negatively with dietary fiber and total carbohydrate content. Additionally, TOS and OSI negatively correlated with parameters related to body weight and positively with insulin. In control men, we observed significant negative correlations between parameters related to fasting glucose and TAS and positive ones with TOS and OSI. CONCLUSIONS: The results show that in lung cancer oxidative stress is related to GI, while TAS is related to nutritional status. Further investigations performed on a larger cohort are required to better clarify the observed relationships as well as to explain the potential mechanisms involved.

The Limitations of Collagen/CPP Hybrid Peptides as Carriers for Cancer Drugs to FaDu Cells.

The in vitro efficacy of cancer prodrugs varies significantly between malignant cell lines. The most commonly identified problems relate to delivery: uptake mechanism, endosomal entrapment, and drug release. Here we present the study of collagen/cell penetrating hybrid (COL/CPP) peptide carriers intended to deliver paclitaxel to the hypopharyngeal carcinoma (FaDu) cells. Confocal microscopy imaging revealed the surprising response of FaDu cell to COL/CPP in comparison to previously studied cancer cell lines: hybrid peptides that carry both COL and CPP domain adsorb on the FaDu cell surface. While the CPP domain was design to facilitate the cellular uptake, in the case of FaDu cells, it also induced detrimental interactions with the cell membrane. Despite surface adsorption, the colocalization study with endosomal markers EEA1 and LAMP1 reveals that COL/CPP is internalized via endosomal pathway, peptides are able to escape before lysosome formation and release paclitaxel. Therefore, the main obstacle for paclitaxel delivery to FaDu cells appears to be related to cell surface properties. This behavior seems specific to FaDu cells, and could be linked to previously reported overexpression of T5, heparanase splice variants that produces protein lacking enzymatic activity of heparanase. This results in increased concentration of HSPG on FaDu cell surface, and possibly creates a barrier for cellular uptake of highly charged COL/CPP.

Occurrence of dietary risk factors in inflammatory bowel disease: Influence on the nutritional status of patients in clinical remission.

BACKGROUND: Among the complex factors that may favor the occurrence of inflammatory bowel disease (IBD), genetic, immunological and environmental initiators, including nutritional factors, are listed. So far, there have been no previous studies on the type and frequency of dietary risk factors for IBD in Poland and their effect on the nutritional status of patients. OBJECTIVES: The aim of the study was to assess the influence of the frequency and type of dietary risk factors for IBD on the nutritional status of patients with ulcerative colitis (UC) and Crohn's disease (CD). MATERIAL AND METHODS: In the study, the dietary habits and nutritional status of patients were assessed using the cross-check dietary history method and the Mini Nutritional Assessment (MNA) questionnaire. The study group consisted of 162 IBD patients: 61 individuals with CD and 101 with UC. The data was compared to the results of a control group (129 healthy volunteers). RESULTS: The results obtained showed that IBD patients during a period of remission disclosed such dietary risk factors as inadequate consumption of fiber and excessive consumption of red meat and meat products, animal fats, and sugars in comparison to the control group. Only low fiber intake was associated with a worse nutritional status of patients with UC. No consistent influence of the number of IBD dietary risk factors on the nutritional status of patients was found. CONCLUSIONS: The nutritional status of IBD patients in remission was related to the type of dietary risk factors, but did not depend on the number of them.

Systemic redox status in lung cancer patients is related to altered glucose metabolism.

Altered systemic redox status is often observed in lung cancer. However, detailed information on factors other, than smoking, which influence this perturbation is rather scarce. Elevated oxidative stress has been linked with disturbances in glucose metabolism before, but such associations have not been investigated in lung cancer. The aim of this study was to evaluate the relationship between systemic parameters of glucose metabolism and redox status in lung cancer patients (LC). Biochemical variables related to circulating glucose, i.e. glucose, insulin, c-peptide, fructosamine (FA), and glucose metabolism, i.e. ß-hydroxybutyrate (BHB), lactate (LACT), non-esterified fatty acids (NEFAs), as well as redox status i.e. total antioxidant status (TAS) and total oxidant status (TOS) were determined for LC (n = 122) and control subjects (CS) (n = 84). HOMA-IR and the oxidative stress index (OSI) were calculated. LC patients had an altered redox status and glucose metabolism compared to CS. Positive correlations in LC were observed between TOS, OSI and circulating glucose as well as FA, while TAS positively correlated with BHB and NEFAs. In contrast, in metastatic LC, NEFAs and BHB positively correlated with OSI. Smoking status additionally stratified the observed relationships. In conclusion, we found that parameters related to circulating glucose or non-enzymatic glycation were correlated with oxidative stress (TOS and OSI), while metabolites such as BHB and NEFAs were correlated with antioxidant capacity (TAS). Metastasis prevalence and smoking seem to influence these correlations. However, the detailed mechanism of this relationship requires further research, in particular as regards the surprising positive correlation between NEFAs and TAS.

The Effect of Pulsed Electric Field on Expression of ECM proteins: Collagen, Elastin, and MMP1 in Human Dermal Fibroblasts.

Electrical stimulation of tissues has many uses in pain management, antibacterial treatment, and wound healing. The electric field stimulates epidermal migration and increases fibroblast cell proliferation. Here we show the effects of electrical field (EF) stimulation of human dermal fibroblasts (HDF) on the expression of collagen, elastin, and collagenase (MMP1; matrix metalloproteinase 1). The effects of EF stimulation are evaluated in terms of changes in cell morphology and extracellular matrix (ECM) protein expression, defined as intracellular concentration of collagen, elastin, and MMP1. HDF are stimulated in a bioreactor using square wave voltage pulses for up to 24 h. The pulse voltage (0-10V), pulse bias (0, +), pulse time (10-1000 ms), and rest time (0.1-10 s) were varied. We show that expression of collagen, elastin, and MMP1 increases in response to applied EF. The intracellular concentration of ECM proteins more than doubles depending on stimulation conditions with a threshold of effective stimulation above 3V/cm. The short time voltage pulses used for EF stimulation are more effective, while the rest time between pulses has a small effect on intracellular concentration of collagen, MMP1 and elastin. The previously studied HDF stimulation with chemical factors (i.e. TNF-α, TGF-ß) shows negative correlation between concentration of collagen and MMP1. Contrary to that observation, we show that EF stimulation causes increase in the intracellular concentration of both collagen and MMP1. We also demonstrate that the transdermal stimulation of HDF in subcutaneous tissue is possible, thus it might be utilized in the future to improve the wound healing and tissue regeneration process.