Pedestrian Safety in Frontal Tram Collision, Part 1: Historical Overview and Experimental-Data-Based Biomechanical Study of Head Clashing in Frontal and Side Impacts.

This article represents the first paper in a two-part series dealing with safety during tram-pedestrian collisions. This research is dedicated to the safety of trams for pedestrians during collisions and is motivated by the increased number of lethal cases. The first part of this paper includes an overview of tram face development from the earliest designs to the current ones in use and, at the same time, provides a synopsis and explanation of the technical context, including a link to current and forthcoming legislation. The historical design development can be characterised by three steps, from an almost vertical front face, to leaned and pointed shapes, to the current inclined low-edged windshield without a protruding coupler. However, since most major manufacturers now export their products worldwide and customisation is only of a technically insignificant nature, our conclusions are generalisable (supported by the example of Berlin). The most advantageous shape of the tram's front, minimising the effects on pedestrians in all collision phases, has evolved rather spontaneously and was unprompted, and it is now being built into the European Commission regulations. The goal of the second part of this paper is to conduct a series of tram-pedestrian collisions with a focus on the frontal and side impacts using a crash test dummy (anthropomorphic test device-ATD). Four tram types approaching the collision at four different impact speeds (5 km/h, 10 km/h, 15 km/h, and 20 km/h) were used. The primary outcome variable was the resultant head acceleration. The risk and severity of possible head injuries were assessed using the head injury criterion (HIC15) and its linkage to the injury level on the Abbreviated Injury Scale (AIS). The results showed increasing head impacts with an increasing speed for all tram types and collision scenarios. Higher values of head acceleration were reached during the frontal impact (17-124 g) compared to the side one (2-84 g). The HIC15 values did not exceed the value of 300 for any experimental setting, and the probability of AIS4+ injuries did not exceed 10%. The outcomes of tram-pedestrian collisions can be influenced by the ATD's position and orientation, the impact speed and front-end design of trams, and the site of initial contact.

Pedestrian Safety in Frontal Tram Collision, Part 2: Laminated Glass as a Crucial Part of the Absorption and Deformation Zone-Its Impact Test and Analysis.

As was shown in the previous part of the study, windshields are an important part of the passive safety means of modern low-floor trams with an extraordinary effect on pedestrian safety in a pedestrian-tram collisions. Therefore, maximum attention must be paid to the definition of tram windshield characteristics. This article describes a windshield crash test, from which data are obtained to verify the feasibility of the applied computational approaches. A developed analytical model is utilised for a simple description of the energy balance during collision with an illustrative definition of the important parameters of laminated glass as well as their clear physical interpretations. The finite element analysis (FEA) performed in Ansys software using two versions of material definition, namely a simpler (*MAT_ELASTIC with nonlocal failure criterion) and a more complex (*MAT_GLASS with brittle stress-state-dependent failure) material model, which are presented as suitable for obtaining a detailed description of the shattering process of laminated glass, which can also be used effectively in windshield engineering.

Load Measurement of the Cervical Vertebra C7 and the Head of Passengers of a Car While Driving across Uneven Terrain.

The article deals with the measurement of dynamic effects that are transmitted to the driver (passenger) when driving in a car over obstacles. The measurements were performed in a real environment on a defined track at different driving speeds and different distributions of obstacles on the road. The reaction of the human organism, respectively the load of the cervical vertebrae and the heads of the driver and passenger, was measured. Experimental measurements were performed for different variants of driving conditions on a 28-year-old and healthy man. The measurement's main objective was to determine the acceleration values of the seats in the vehicle in the vertical movement of parts of the vehicle cabin and to determine the dynamic effects that are transmitted to the driver and passenger in a car when driving over obstacles. The measurements were performed in a real environment on a defined track at various driving speeds and diverse distributions of obstacles on the road. The acceleration values on the vehicle's axles and the structure of the driver's and front passenger's seats, under the buttocks, at the top of the head (Vertex Parietal Bone) and the C7 cervical vertebra (Vertebra Cervicales), were measured. The result of the experiment was to determine the maximum magnitudes of acceleration in the vertical direction on the body of the driver and the passenger of the vehicle when passing a passenger vehicle over obstacles. The analysis of the experiment's results is the basis for determining the future direction of the research.

Fallacies and Pitfalls of Dialysis Sodium Prescription and Control.

Dialysate conductivity is routinely used as a surrogate for dialysate sodium concentration. However, dialysis machine manufacturers apply different conductivity temperature correction coefficients. With the same conductivity in dialysis machines manufactured by different manufacturers, dialysate sodium may significantly differ. Also, electrolyte prescriptions are individualized (K, Ca, HCO3) and this is associated with another variation in dialysate sodium in the order of 1-5 mmol/L and both deviations are cumulative and chronic for each patient. Equivalence of the prescribed dialysate sodium and the concentration measured in it is not granted. Both variables differ and it is machine dependent! This paper analyses those variations from a technical point of view and suggests how to detect them and how to deal with or avoid them in clinical practice.

Calcaneal Tendon Collagen Fiber Morphometry and Aging.

Fibrillar collagen in tendons and its natural development in rabbits are discussed in this paper. Achilles tendons from newborn (~7 days) to elderly (~38 months) rabbits were monitored in intact (n tendons=24) and microtome sectioned (n tendons=11) states with label-free second harmonic generation microscopy. After sectioning, the collagen fiber pattern was irregular for the younger animals and remained oriented parallel to the load axis of the tendon for the older animals. In contrast, the collagen fiber pattern in the intact samples followed the load axis for all the age groups. However, there was a significant difference in the tendon crimp pattern appearance between the age groups. The crimp amplitude (A) and wavelength (Λ) started at very low values (A=2.0±0.6 µm, Λ=19±4 µm) for the newborn animals. Both parameters increased for the sexually mature animals (>5 months old). When the animals were fully mature the amplitude decreased but the wavelength kept increasing. The results revealed that the microtome sectioning artifacts depend on the age of animals and that the collagen crimp pattern reflects the physical growth and development.

Evaluation of rheological parameters of the axial system using the Transfer Vibration through Spine (TVS) method.

OBJECTIVE: The human motion system reacts to both hypo and hyperactivity loads by changes to the rheological properties of tissues. This study deals with changes to the axial system (AS) compartment. Using the appropriate methodologies and mathematical-physical methods, these changes can be identified and quantified. METHODS: This study describes the noninvasive TVS (Transfer Vibration through Spine) method, which was applied to assess the AS selected mechanical properties in various modes. A pilot study was conducted on a top-level twelve-year-old girl-gymnast. The data detection was carried out in three cycles, before and after a peak 3.5 hour training session and the next day, after resting, just before the next training. RESULTS: Specifically, the values of selected rheological parameters, the AS damping coefficient b and viscosity µ, were obtained. The dynamics of their changes, in the stated load cycles, has also been shown. The damping coefficient b fell from the value of 0.626 to 0.324 before training and increased to 0.394 after resting. The viscosity coefficient µ showed a similar trend, namely falling from the value of 9.85 [Pa.s] to 2.15 [Pa.s] and then increasing to 3.8 [Pa.s] . CONCLUSIONS: With its computational solution, the TVS method,is a diagnostic apparatus making it possible to classify AS properties, both quantitatively and qualitatively, or its chosen segments and their changes, respectively. It can be used in classifying, preventing and treating the consequences of extreme motion and relaxing modes. The TVS application also makes it possible to control AS states over therapeutic, recovery, ergonomic and other loading modes of the human locomotion system.

[Bioimpedometry and its utilization in dialysis therapy]. / Bioimpedometrie a její vyuzití v dialyzacní lécbe.

Measurement of living tissue impedance - bioimpedometry - started to be used in medicine some 50 years ago, first exclusively for estimation of extracellular and intracellular compartment volumes. Its most simple single frequency (50 kHz) version works directly with the measured impedance vector. Technically more sophisticated versions convert the measured impedance in values of volumes of different compartments of body fluids and calculate also principal markers of nutritional status (lean body mass, adipose tissue mass). The latest version specifically developed for application in dialysis patients includes body composition modelling and provides even absolute value of overhydration (excess fluid). Still in experimental phase is the bioimpedance exploitation for more precise estimation of residual glomerular filtration. Not yet standardized is also segmental bioimpedance measurement which should enable separate assessment of hydration status of the trunk segment and ultrafiltration capacity of peritoneum in peritoneal dialysis patients.Key words: assessment - bioimpedance - excess fluid - fluid status - glomerular filtration - haemodialysis - nutritional status - peritoneal dialysis.

Pre-activation and muscle activity during frontal impact in relation to whiplash associated disorders.

OBJECTIVE: For the evaluation of neck injury the relative distance was observed between a marker placed on the forehead and a marker placed on the shoulder and also by change of the angle. To compare the severity of head injury a value of maximum head acceleration was used, HIC and a 3 ms criterion. All criteria were related to the activity of musculus sternocleidomastoideus and musculus trapezius in a situation of expected or unexpected impact. MATERIALS AND METHODS: The situation was recorded using a Qualisys system, head acceleration of probands in three axes was recorded using the accelerometer, activity of neck muscles was monitored by a mobile EMG. RESULTS: Maximum head acceleration was 12.1 g for non-visual and 8.2 g for visual. HIC36 was 5.7 non visual and 4.0 for visual. 3-ms criterion was 11.5 g for non-visual and 7.8 g for visual. The average time of muscle activation of the observed group without visual perception is 0.027 s after hitting an obstacle, with visual perception 0.127 s before the crash. CONCLUSIONS: Kinematic values indicate more favourable parameters for neck injuries for visual. Head injury criteria show an average decrease of about 30% for visual. We can conclude that the visual perception means a significant increase in pre-activation of the observed muscle group of almost 400% and lower activation in both following phases of approximately 40%.

Intervertebral (lumbar) disc replacement:the current state and future perspectives.

PURPOSE: Low back pain is a significant socio-economic problem which is expected to deepen. Degenerative disc disease is considered to be one of its main causes. Unsuccessful conservative treatments usually lead to surgical treatments, including methods providing pain relief by vertebral fusion in the affected segment. However, this leads to changes in biomechanics, which is why approximately 30 years ago total disc replacements appeared. This work aims at determining the current state of treatments with this kind of replacement, comparing the results with those of fusion methods and assessing why fusion continues to dominate. Current treatments of degenerative disc disease by advanced procedures (regenerative and gene therapy, 3D printing) is also examined and evaluated, and future developments are considered. METHODS: A critical review based on available scientific articles from online databases. The main keywords used were "lumbar", "total", "disc" and "replacement", supplemented according to the individual, monitored areas ("follow-up", "fusion", "future" etc.). For the articles found through database search (n = 895), narrower selection was made and the result was 33 articles included in review. REVIEW: Total disc replacements have not yet satisfactorily demonstrated that they are superior to fusion methods in long term follow-up. Advanced methods are in their infancy. CONCLUSIONS: Additional research and development of total disc replacements is still necessary. For implants, the 3D scan - 3D model - 3D printing chain and its related technologies are increasingly important. The development of regenerative procedures using induced pluripotent stem cells and gene therapies is important, but conservative treatments and primary prevention should also be developed because regenerative procedures and gene therapies apparently will not be used routinely until the future.

The Incidence of Clubfoot in the Czech Republic: A Nationwide Epidemiological Study from 2000 to 2014.

Clubfoot is one of the most common musculoskeletal birth deformities worldwide. The prevalence varies among individual countries and populations. There is a lack of nationwide incidence studies in Central Europe. We analyzed the incidence of clubfoot in the Czech Republic over 14 years. Patients born with clubfoot in the Czech Republic were identified using The National Registry of Congenital Anomalies. Demographic data were included. Data from 2000 to 2014 were collected and analyzed regarding gender and regional distribution. The study's chosen time frame was grounded on the condition of the Czech industry. Following extensive transformations in 1989, the industry eliminated highly non-ecological operations with significant environmental impact and related health risks. The incidence of clubfoot during the study period was 1.9 (95% CI 1.8-2.0) per 1000 births; males comprised the majority (59%). The incidence significantly differed among individual regions of the Czech Republic (p < 0.001). The incidence in the Czech Republic was higher than in previous European studies. We found significant regional differences in incidence, which could indicate that there may be exogenous pathogenic factors. For this reason, we plan to follow up our work with an up-to-date study.

Influence of SYSADOA group chemicals on progression of human knee joint osteoarthritis: new objective evaluation method - measuring of rheological properties in vivo.

OBJECTIVES: This study seeks to demonstrate the influence of pharmacological substances from the SYSADOA group on the progression of osteoarthritis in the human knee. The quantification methods were direct measurement of the rheological properties of the knee joints in vivo and standard WOMAC index questionnaires. MATERIALS AND METHODS: The drugs were administered orally to 34 probands with second degree gonarthrosis for 13 weeks. The untreated control group consisted of 10 probands. The rheological properties of the joints were determined by a biorheometer, and subjective assessment of the knees by patients (WOMAC) before and after medication, and for a further 13 weeks. Changes in the calculated parameters over time were compared. RESULTS: During the audited perioda slight deterioration in all of the parameters was observed in the untreated group. The treated group, however, improved in all the parameters and some indicators showed statistically significant differences. The positive effects of the SYSADOA persisted for 3 months after the end of treatments. Partial correlation was found between the results of the WOMAC questionnaire and the rheological measurements. CONCLUSIONS: This study shows the positive effects of the preparation on arthritic changes in the knee joint, but due to the large variance of the collected data, this conclusion is on the borderline of statistical significance. The method of measuring the rheological properties of the joints is suitable for evaluating the progression of OA.

The ERA Registry Annual Report 2019: summary and age comparisons.

BACKGROUND: Data on renal replacement therapy (RRT) for end-stage renal disease were collected by the European Renal Association (ERA) Registry via national and regional renal registries in Europe and countries bordering the Mediterranean Sea. This article provides a summary of the 2019 ERA Registry Annual Report, including data from 34 countries and additional age comparisons. METHODS: Individual patient data for 2019 were provided by 35 registries and aggregated data by 17 registries. Using these data, the incidence and prevalence of RRT, the kidney transplantation activity and the survival probabilities were calculated. RESULTS: In 2019, a general population of 680.8 million people was covered by the ERA Registry. Overall, the incidence of RRT was 132 per million population (p.m.p.). Of these patients, 62% were men, 54% were ≥65 years of age and 21% had diabetes mellitus as primary renal disease (PRD), and 84% had haemodialysis (HD), 11% had peritoneal dialysis (PD) and 5% had pre-emptive kidney transplantation as an initial treatment modality. The overall prevalence of RRT on 31 December 2019 was 893 p.m.p., with 58% of patients on HD, 5% on PD and 37% living with a kidney transplant. The overall kidney transplant rate was 35 p.m.p. and 29% of the kidney grafts were from a living donor. The unadjusted 5-year survival probability was 42.3% for patients commencing dialysis, 86.6% for recipients of deceased donor grafts and 94.4% for recipients of living donor grafts in the period 2010-14. When comparing age categories, there were substantial differences in the distribution of PRD, treatment modality and kidney donor type, and in the survival probabilities.

Changes in vascular access blood flow: Etiological factors and clinical implications.

METHODS: Records of 10,000 QVA measurement performed in 549 patients over 20 years were used as retrospective and anonymized data source, making ethical commission involvement unnecessary. Two approaches are used to elucidate association of QVA changes with different factors: analyses of smaller cohorts in which both the QVA and the respective factor were measured (e.g. association of QVA with cardiac output (CO)), or-in case of rare phenomena-a form of a well illustrated case reports was used (e.g. association of QVA and Kt/V). RESULTS: Significant increase in CO after permanent VA creation (3-4-fold of the QVA value) was found. Impact of intradialytic CO changes on QVA is attenuated by relatively stable VA resistance compared to systemic resistance. Blood pressure impact is much stronger and it should therefore be noted at each QVA measurement. As reproducibility of different QVA measurement methods varies, use of the same method should be preferred. Direction of the arterial needle insertion in VA affects the QVA measured, especially in synthetic grafts, too. Also patient's own QVA variability may be quite high. All this makes KDOQI/EBPG recommended acceptable QVA drops too strict, they should be revised. In re-stenoses prone patients, measurement intervals should be shortened, too. CONCLUSION: QVA values are significantly affected by many factors. Their knowledge appears essential for safe and effective VA surveillance and management.

The ERA-EDTA Registry Annual Report 2018: a summary.

BACKGROUND: The European Renal Association - European Dialysis and Transplant Association (ERA-EDTA) Registry collects data on kidney replacement therapy (KRT) via national and regional renal registries in Europe and countries bordering the Mediterranean Sea. This article summarizes the 2018 ERA-EDTA Registry Annual Report, and describes the epidemiology of KRT for kidney failure in 34 countries. METHODS: Individual patient data on patients undergoing KRT in 2018 were provided by 34 national or regional renal registries and aggregated data by 17 registries. The incidence and prevalence of KRT, the kidney transplantation activity and the survival probabilities of these patients were calculated. RESULTS: In 2018, the ERA-EDTA Registry covered a general population of 636 million people. Overall, the incidence of KRT for kidney failure was 129 per million population (p.m.p.), 62% of patients were men, 51% were ≥65 years of age and 20% had diabetes mellitus as cause of kidney failure. Treatment modality at the onset of KRT was haemodialysis (HD) for 84%, peritoneal dialysis (PD) for 11% and pre-emptive kidney transplantation for 5% of patients. On 31 December 2018, the prevalence of KRT was 897 p.m.p., with 57% of patients on HD, 5% on PD and 38% living with a kidney transplant. The transplant rate in 2018 was 35 p.m.p.: 68% received a kidney from a deceased donor, 30% from a living donor and for 2% the donor source was unknown. For patients commencing dialysis during 2009-13, the unadjusted 5-year survival probability was 42.6%. For patients receiving a kidney transplant within this period, the unadjusted 5-year survival probability was 86.6% for recipients of deceased donor grafts and 93.9% for recipients of living donor grafts.

Women with incorrect pelvic floor statics: a biomechanical answer to the mechanical loading of the vagina-endopelvic fascia complex.

OBJECTIVE: This work focuses on finding method for detecting the elementary mechanical characteristics of the vagina-endopelvic fascia complex, aimed at providing results for use in optimizing solutions for stability defects of the pelvic floor. MATERIALS AND METHODS: Two experiments have already been carried out that have enabled monitoring of the reaction of tissue complex samples to a selected load. The elastic properties of samples under a simple pull load were evaluated. The monitored property in the first experiment was the maximum reference tension at the moment of rupture of the sample in relation to the non-deformed section. We evaluated data from measurements on 11 samples within the scope of the first experiment. For data processing from the second experiment we used a linear-elastic model of the sample, formed by parallel connection of basic mechanical elements - springs - that represented the endopelvic fascia and the vaginal wall. The relevant rigidities were used for a description of their properties. Five samples were used for this experiment. RESULTS: An important discovery was that the endopelvic fascia tears apart after a longer period of time than the vaginal wall during the pull test. The results show considerable variability among individuals, but the pattern of curves is similar in all test cases. In all measured data we found a rigidity increase zone, a maximum rigidity zone and a gradual rigidity decrease zone before terminal damage in the response. CONCLUSIONS: The results presented here show quite broad interindividual variability of the mechanical properties of the vaginal wall-endopelvic fascia complex. It appears that the mechanical properties of the tissue complex change with number of pregnancies, and are affected by diseases, by physical load or by the presence of other factors, e.g. obesity.

The ERA-EDTA Registry Annual Report 2017: a summary.

BACKGROUND: This article presents a summary of the 2017 Annual Report of the European Renal Association-European Dialysis and Transplant Association (ERA-EDTA) Registry and describes the epidemiology of renal replacement therapy (RRT) for end-stage renal disease (ESRD) in 37 countries. METHODS: The ERA-EDTA Registry received individual patient data on patients undergoing RRT for ESRD in 2017 from 32 national or regional renal registries and aggregated data from 21 registries. The incidence and prevalence of RRT, kidney transplantation activity and survival probabilities of these patients were calculated. RESULTS: In 2017, the ERA-EDTA Registry covered a general population of 694 million people. The incidence of RRT for ESRD was 127 per million population (pmp), ranging from 37 pmp in Ukraine to 252 pmp in Greece. A total of 62% of patients were men, 52% were ≥65 years of age and 23% had diabetes mellitus as the primary renal disease. The treatment modality at the onset of RRT was haemodialysis for 85% of patients. On 31 December 2017, the prevalence of RRT was 854 pmp, ranging from 210 pmp in Ukraine to 1965 pmp in Portugal. The transplant rate in 2017 was 33 pmp, ranging from 3 pmp in Ukraine to 103 pmp in the Spanish region of Catalonia. For patients commencing RRT during 2008-12, the unadjusted 5-year patient survival probability for all RRT modalities combined was 50.8%.

Obliteration Index: a simple non-invasive tool for the assessment of peripheral vascular disease in chronic kidney disease.

BACKGROUND: Peripheral artery disease (PAD) represents a major complication in chronic kidney disease (CKD) patients, but its early diagnosis is still problematic. Traditionally, ankle-arm blood pressure index (API) determined by means of a simple hand-held unidimensional Doppler device has been used for noninvasive diagnosis of PAD, with a cutoff value of 0.9. However, in CKD patients, vascular disease includes not only atherosclerosis but also arteriosclerosis with high calcification score. There, the API method often fails. DEVELOPMENT OF THE METHOD: Based on complex examination of Doppler signals recorded at 4 main arteries of both lower extremities (a. femoralis, a. poplitea, a. dorsalis pedis, a. tibialis posterior) and their semiquantitative assessment, we introduced the Obliteration Index (OI) as a novel and simple tool allowing semiquantitative assessment of PAD in CKD patients: the signal from each artery is assessed in a 6-grade scale, with each grade represented by 5 points. The sum of points allocated to all arteries constitutes the final OI value, ranging from 0 (normal finding) up to 240 (most severe PAD). VERIFICATION OF THE METHOD: Comparison of the OI method with the conventional API examined in over 100 renal patients revealed superior OI sensitivity in detection of PAD in the early stage and low-grade vessel infliction where the API still remains within the normal range. OI can also be evaluated even when API is not measurable. Very good correspondence was found between the OI and angiography. CONCLUSIONS: The OI method has now been introduced into routine use in the authors' department. Based on over 1,500 OI evaluations performed so far, it can be recommended as a simple, noninvasive, bedside tool for diagnosis of early as well as advanced PAD.

Improved intradialytic stability during haemodialysis with blood volume-controlled ultrafiltration.

BACKGROUND: Intradialytic morbid events (IMEs) during haemodialysis (HD), including symptomatic hypotension, are related to ultrafiltration (UF)-induced hypovolaemia. Blood volume monitoring and automatic feedback control of the UF rate were developed to limit the extent of hypovolaemia during dialysis. The present study investigated the effect of blood volume (BV)-controlled UF on the incidence of HD treatments with IMEs. METHODS: This prospective randomised crossover study included hypotension-prone patients, characterised by occurrence of IMEs in at least 33% of HD treatments during a 6-week screening phase. These patients underwent 2 treatment phases, each lasting 6 weeks, in randomised order. Each patient served as their own control, treated with standard HD in one phase and with BV-controlled UF in the other phase. RESULTS: Thirty-four patients from 9 HD centres were enrolled; 26 could be included in the analysis population. In comparison with standard HD, BV-controlled UF reduced the percentage of HD sessions complicated by IME significantly from 40%+/-27% to 32%+/-25% (p=0.02). A lower frequency of HD sessions with IME could be observed in 46% of the patients. The frequency of treatments with symptomatic hypotension was reduced from 32%+/-23% in standard HD to 24%+/-21% with BV-controlled UF (p=0.04). Changes in blood pressure and heart rate from start to end of the HD session were not different between the 2 treatment modes. CONCLUSIONS: This crossover study showed improved intradialytic stability with BV-controlled UF, compared with standard HD.

A two-layer skin construct consisting of a collagen hydrogel reinforced by a fibrin-coated polylactide nanofibrous membrane.

Background: Repairs to deep skin wounds continue to be a difficult issue in clinical practice. A promising approach is to fabricate full-thickness skin substitutes with functions closely similar to those of the natural tissue. For many years, a three-dimensional (3D) collagen hydrogel has been considered to provide a physiological 3D environment for co-cultivation of skin fibroblasts and keratinocytes. This collagen hydrogel is frequently used for fabricating tissue-engineered skin analogues with fibroblasts embedded inside the hydrogel and keratinocytes cultivated on its surface. Despite its unique biological properties, the collagen hydrogel has insufficient stiffness, with a tendency to collapse under the traction forces generated by the embedded cells. Methods: The aim of our study was to develop a two-layer skin construct consisting of a collagen hydrogel reinforced by a nanofibrous poly-L-lactide (PLLA) membrane pre-seeded with fibroblasts. The attractiveness of the membrane for dermal fibroblasts was enhanced by coating it with a thin nanofibrous fibrin mesh. Results: The fibrin mesh promoted the adhesion, proliferation and migration of the fibroblasts upwards into the collagen hydrogel. Moreover, the fibroblasts spontaneously migrating into the collagen hydrogel showed a lower tendency to contract and shrink the hydrogel by their traction forces. The surface of the collagen was seeded with human dermal keratinocytes. The keratinocytes were able to form a basal layer of highly mitotically-active cells, and a suprabasal layer. Conclusion: The two-layer skin construct based on collagen hydrogel with spontaneously immigrated fibroblasts and reinforced by a fibrin-coated nanofibrous membrane seems to be promising for the construction of full-thickness skin substitute.