Fibroblast Density Research Articles

EFFECT OF LOCAL UV RADIATION ON THE MORPHOFUNCTIONAL STATE OF THE SKIN OF GUINEA PIGS IN EARLY AND REMOTE PERIODS AFTER IRRADIATION

The aim is to establish the features of morphological and morphometric changes in the skin of guinea pigs in erythemal, early post-erythemal and late post-erythemic periods after local ultraviolet irradiation. Materials and methods: Studies were conducted on 54 albino guinea pigs weighing 400-500 g. Ultraviolet erythema was caused by irradiation in 1 minimum erythemal dose. The control group included intact guinea pigs. After 2, 4 hours, on the 3rd, 8th, 15th, 21st, 28th day, the fragments of the irradiated skin were investigated using histochemical and morphometric methods. Results: After 2, 4 hours after irradiation, dyscirculatory changes in the skin develop. By the 3rd day of the experiment a morphological picture of acute inflammation in the epidermis and dermis develops, apoptotic keratinocytes appear (sunburn cells), which is accompanied by thickening of the epidermis and an increase in the density of fibroblasts. By the 8th day proliferative-hyperplastic and degenerative changes begin to prevail, including dystrophic nature, the thickness of the epidermis and the density of fibroblasts reach a maximum. In the long term, on the 15-28th day, dystrophic changes of the epidermis, dyskeratosis, changes in the number and structure of elastic fibers with an increase in uneven fibrosis, collagenization processes and the development of sclerotic changes, as well as a significant thickening of the epidermis, an increase in the density of fibroblasts are observed. Conclusions: The data obtained indicate pronounced morphofunctional changes in the skin in the zone of local ultraviolet irradiation observed throughout the entire observation period.

THE EFFECT OF OINTMENTS WITH ANTIOXIDANT ACTIVITY ON THE MORPHOLOGICAL STRUCTURES OF SKIN OF GUINEA PIGS EXPOSED TO LOCAL ULTRAVIOLET IRRADIATION

Considering the particular danger of remote skin reactions to ultraviolet irradiation (UVI), it is advisable to use ointments with antioxidant activity to reduce its negative effect on the skin. The rationale for the choice of ointments with antioxidant activity was the fact that they reduce the damaging effect of ultraviolet radiation in the erythemal and early post-erythemal period. The presence of a regular connection between the development of the early and late periods has given reason to assume the protective effect of ointments on the remote skin reactions.
 Objective: to study the effect of thiotriazoline ointment and thiotriazoline ointment with silver nanoparticles on the state of the morphological structures of the skin of guinea pigs after local UVI.
 Material and methods of research. The study involved 132 albino guinea pigs weighing 400-500 g, divided into 4 groups: 1 - intact, 2 - control (guinea pigs subjected to local UVI), 3 and 4 main ones. The third main group included guinea pigs that after UVI were administered thiotriazoline ointment in the treatment and prophylactic regime, the fourth main group included guinea pigs that after UVI were administered thiotriazoline ointment with silver nanoparticles in the same mode as Group 3. Ointments were applied 1 hour before irradiation and daily until erythema disappeared. Ultraviolet erythema was caused by irradiation in 1 minimum erythemal dose. After 2, 4 hours, on the 3rd, 8th, 15th, 21st, 28th day, the fragments of irradiated skin were investigated using histochemical and morphometric methods (fibroblast density and epidermis thickness).
 Results. Morphological changes in the skin after applying ointments with antioxidant activity were unidirectional. It was revealed that in the early periods after irradiation, thiotrazoline ointment and thiotrazoline ointment with silver nanoparticles do not affect changes in the thickness of the epidermis, but statistically significantly reduce the density of fibroblasts in the dermis on the 3rd day of the experiment compared to the control group.
 In the later periods, under the influence of thiotriazoline ointment, a gradual decrease in the thickness of the epidermis, which reached the norm by the end of the experiment, was observed. On the 8th day, the maximum density of fibroblasts was recorded, in the subsequent periods of the experiment, the index gradually decreased, which was accompanied by collagenization of the papillary layer in the loci of damage to collagen and elastic fibers detected in 50% of cases.
 In later times, under the influence of thiotriazoline ointment with silver nanoparticles, the processes of restoring the morphological structures of the skin occurred faster. In parallel with the decrease in the density of fibroblasts in the loci of the previous damage to the collagen and elastic fibers of the papillary layer, thickening of collagen fibers was observed, replacing them with segments of destruction of elastic fibers. In this group, at the end of the experiment, the collagenization locus was small, single, occurring in 16.7% of cases.
 Conclusions
 
 Ointments with antioxidant activity exert a positive effect on the state of morphological structures of the skin, damaged as a result of local UVI, in erythemal and post-erythemic periods.
 In the early periods after the local UVI, there was a general tendency for the effect of both ointments, as they reduced the density of fibroblasts on the 3rd day, but did not result in complete normalization.
 In the late period after local UVI , under the influence of thiotriazoline ointment and thiotriazoline ointment with silver nanoparticles, thickness of the epidermis (by 21st and 15th day, respectively) and density of fibroblasts (by the 28th day) decreased to normal while without treatment both indicators exceeded the norm by several times for 28 days of the experiment.

THE EFFECT OF OINTMENTS WITH ANTIOXIDANT ACTIVITY ON THE MORPHOLOGICAL STRUCTURES OF SKIN OF GUINEA PIGS EXPOSED TO LOCAL ULTRAVIOLET IRRADIATION

Considering the particular danger of remote skin reactions to ultraviolet irradiation (UVI), it is advisable to use ointments with antioxidant activity to reduce its negative effect on the skin. The rationale for the choice of ointments with antioxidant activity was the fact that they reduce the damaging effect of ultraviolet radiation in the erythemal and early post-erythemal period. The presence of a regular connection between the development of the early and late periods has given reason to assume the protective effect of ointments on the remote skin reactions.
 Objective: to study the effect of thiotriazoline ointment and thiotriazoline ointment with silver nanoparticles on the state of the morphological structures of the skin of guinea pigs after local UVI.
 Material and methods of research. The study involved 132 albino guinea pigs weighing 400-500 g, divided into 4 groups: 1 - intact, 2 - control (guinea pigs subjected to local UVI), 3 and 4 main ones. The third main group included guinea pigs that after UVI were administered thiotriazoline ointment in the treatment and prophylactic regime, the fourth main group included guinea pigs that after UVI were administered thiotriazoline ointment with silver nanoparticles in the same mode as Group 3. Ointments were applied 1 hour before irradiation and daily until erythema disappeared. Ultraviolet erythema was caused by irradiation in 1 minimum erythemal dose. After 2, 4 hours, on the 3rd, 8th, 15th, 21st, 28th day, the fragments of irradiated skin were investigated using histochemical and morphometric methods (fibroblast density and epidermis thickness).
 Results. Morphological changes in the skin after applying ointments with antioxidant activity were unidirectional. It was revealed that in the early periods after irradiation, thiotrazoline ointment and thiotrazoline ointment with silver nanoparticles do not affect changes in the thickness of the epidermis, but statistically significantly reduce the density of fibroblasts in the dermis on the 3rd day of the experiment compared to the control group.
 In the later periods, under the influence of thiotriazoline ointment, a gradual decrease in the thickness of the epidermis, which reached the norm by the end of the experiment, was observed. On the 8th day, the maximum density of fibroblasts was recorded, in the subsequent periods of the experiment, the index gradually decreased, which was accompanied by collagenization of the papillary layer in the loci of damage to collagen and elastic fibers detected in 50% of cases.
 In later times, under the influence of thiotriazoline ointment with silver nanoparticles, the processes of restoring the morphological structures of the skin occurred faster. In parallel with the decrease in the density of fibroblasts in the loci of the previous damage to the collagen and elastic fibers of the papillary layer, thickening of collagen fibers was observed, replacing them with segments of destruction of elastic fibers. In this group, at the end of the experiment, the collagenization locus was small, single, occurring in 16.7% of cases.
 Conclusions
 
 Ointments with antioxidant activity exert a positive effect on the state of morphological structures of the skin, damaged as a result of local UVI, in erythemal and post-erythemic periods.
 In the early periods after the local UVI, there was a general tendency for the effect of both ointments, as they reduced the density of fibroblasts on the 3rd day, but did not result in complete normalization.
 In the late period after local UVI , under the influence of thiotriazoline ointment and thiotriazoline ointment with silver nanoparticles, thickness of the epidermis (by 21st and 15th day, respectively) and density of fibroblasts (by the 28th day) decreased to normal while without treatment both indicators exceeded the norm by several times for 28 days of the experiment.

MORPHOLOGICAL ASSESSMENT OF STRUCTURAL CHANGES IN KNEE JOIN IN MODELED OSTEOARTHROSIS AND APPLICATION OF AUTOLOGICAL BONE MARROW ASPIRATE

Background. Treatment of knee join osteoarthritis is one of challenging medico-social issues. Sometimes the conservative treatment fails to produce therapeutic effect, therefore, the use of cellular technologies in the treatment for progressive degenerative changes of the knee joint has been proposed. The objective of this study was to investigate the effect of bone marrow aspirate on the structural changes in the knee joint in modelled osteoarthritis. Materials and methods. The experimental osteoarthritis was simulated in rabbits. Autologous bone marrow aspirate was injected into the knee cavity in 1 month after surgery, and then in 2, 5 months the structural changes of the capsule, meniscus, proximal epiphysis of the tibia were studied. Results. There were the following structural changes in the capsule of the knee joint: stratification of structural elements and reduction of the cell density of the capsule, cell necrosis of the inner surface of the capsule. The defect area of the epiphyseal cartilage was lacking in cartilage, and the peripheral cartilage thickness was decreased by 30, 4% (p<0,05); subchondral bone density made up an average 55,9% compared with the control. In the group with bone marrow aspirate, the density of fibroblasts in the joint capsule increased and the surface of the meniscus was partially preserved; the density of subchondral bone tissue was significantly higher by 79,5% (p<0.05) compared with the control. Structural changes in osteoarthritis include progressive dystrophic changes of the joint surface, subchondral bone, meniscus, and joint capsule. The use of autologous bone marrow aspirate can prevent damage to the subchondral bone, meniscus and joint capsule and promote reticular tissue formation and induction of angiogenesis. The morphological changes of bone marrow through progressive osteoarthritis and its renovation after local application of autological bone marrow aspirate concentrate are promising approaches for the further studies.

Effects of phenytoin injection on vocal cord healing after mechanical trauma: An experimental study

Background/aimPhenytoin is an anticonvulsant drug which causes fibroblast proliferation, collagen synthesis, and an increase in epidermal growth factor. Therefore, the aim of the present study is to evaluate the effect of phenytoin injection on the wound healing process in rats with vocal cord injury by histopathological methods.Materials and methodsThe vocal cords of 10 albino Wistar rats were damaged bilaterally; the left vocal cord was kept as the control group. Phenytoin was injected in the right vocal cord. Ten rats were sacrificed. The thickness of the lamina propria and density of the fibroblast and collagen were evaluated histopathologically.ResultsThickness of the lamina propria was 18.0 ± 7.1 µm in the control group, 65.5 ± 10.7 µm in the phenytoin group. The density of fibroblast and collagen were statistically lower in the control group compared the phenytoin group (P < 0.05).ConclusionPhenytoin injection in rats after vocal cord injury significantly increased the thickness of the lamina propria and density of fibroblast and regular and mature collagen in the lamina propria. The findings in our study provide a feasible scientific view for adding phenytoin treatment to vocal cord surgeries in otolaryngology practice, but further studies are needed in order to evaluate the use of phenytoin in preventing the formation of scar tissue and possible effects on vocal cord vibration in humans after vocal cord injury.

Impact of Extravasated Platelet Activation and Podoplanin-positive Cancer-associated Fibroblasts in Pancreatic Cancer Stroma.

The aim of the study was to evaluate the status of extravasated platelet activation (EPA) surrounding podoplanin (PDPN)-positive cancer-associated fibroblasts (CAFs) in pancreatic cancer stroma by neoadjuvant chemotherapy. A total of 74 patients were enrolled in this study. We investigated CD42b and PDPN expression in the groups of untreated, gemcitabine (GEM) alone, GEM plus S-1 (GS) and GEM plus nab-paclitaxel (GnP). CD42b expression in surrounding CAFs was observed in 58% patients. CD42b expression was significantly correlated with PDPN expression. CD42b-positive cases were significantly lower in the group treated with GnP than in the untreated group and groups treated with GEM alone or GS. PDPN expression was reduced in the GnP group, as revealed by markedly disorganized collagen and a low density of PDPN-positive fibroblasts. There was a significantly lower CD42b expression and fewer PDPN-positive fibroblasts in the GnP group than in untreated, GEM alone, and GS groups, but there was no significant difference between the latter three groups. There is a significant association between EPA and PDPN-positive CAFs in pancreatic cancer stroma. Our data suggest that the GnP regimen decreases EPA through PDPN-positive CAF depletion.

Molecular Imaging of Fibroblast Activity After Myocardial Infarction Using a 68Ga-Labeled Fibroblast Activation Protein Inhibitor, FAPI-04.

Heart failure remains a major source of late morbidity and mortality after myocardial infarction (MI). The temporospatial presence of activated fibroblasts in the injured myocardium predicts the quality of cardiac remodeling after MI. Therefore, monitoring of activated fibroblasts is of great interest for studying cardiac remodeling after MI. Fibroblast activation protein (FAP) expression is upregulated in activated fibroblasts. This study investigated the feasibility of imaging activated fibroblasts with a new 68Ga-labeled FAP inhibitor (68Ga-FAPI-04) for PET imaging of fibroblast activation in a preclinical model of MI. Methods: MI and sham-operated rats were scanned with 68Ga-FAPI-04 PET/CT (1, 3, 6, 14, 23, and 30 d after MI) and with 18F-FDG (3 d after MI). Dynamic 68Ga-FAPI-04 PET and blocking studies were performed on MI rats 7 d after coronary ligation. After in vivo scans, the animals were euthanized and their hearts harvested for ex vivo analyses. Cryosections were prepared for autoradiography, hematoxylin and eosin (H&E), and immunofluorescence staining. Results:68Ga-FAPI-04 uptake in the injured myocardium peaked on day 6 after coronary ligation. The tracer accumulated intensely in the MI territory, as identified by decreased 18F-FDG uptake and confirmed by PET/MR and H&E staining. Autoradiography and H&E staining of cross-sections revealed that 68Ga-FAPI-04 accumulated mainly at the border zone of the infarcted myocardium. In contrast, there was only minimal uptake in the infarct of the blocked rats, comparable to the uptake in the remote noninfarcted myocardium (PET image-derived ratio of infarct uptake to remote uptake: 6 ± 2). Immunofluorescence staining confirmed the presence of FAP-positive myofibroblasts in the injured myocardium. Morphometric analysis of the whole-heart sections demonstrated 3- and 8-fold higher FAP-positive fibroblast density in the border zone than in the infarct center and remote area, respectively. Conclusion:68Ga-FAPI-04 represents a promising radiotracer for in vivo imaging of post-MI fibroblast activation. Noninvasive imaging of activated fibroblasts may have significant diagnostic and prognostic value, which could aid clinical management of patients after MI.

Correlation Analysis of Histopathology and Proteogenomics Data for Breast Cancer

Tumors are heterogeneous tissues with different types of cells such as cancer cells, fibroblasts, and lymphocytes. Although the morphological features of tumors are critical for cancer diagnosis and prognosis, the underlying molecular events and genes for tumor morphology are far from being clear. With the advancement in computational pathology and accumulation of large amount of cancer samples with matched molecular and histopathology data, researchers can carry out integrative analysis to investigate this issue. In this study, we systematically examine the relationships between morphological features and various molecular data in breast cancers. Specifically, we identified 73 breast cancer patients from the TCGA and CPTAC projects matched whole slide images, RNA-seq, and proteomic data. By calculating 100 different morphological features and correlating them with the transcriptomic and proteomic data, we inferred four major biological processes associated with various interpretable morphological features. These processes include metabolism, cell cycle, immune response, and extracellular matrix development, which are all hallmarks of cancers and the associated morphological features are related to area, density, and shapes of epithelial cells, fibroblasts, and lymphocytes. In addition, protein specific biological processes were inferred solely from proteomic data, suggesting the importance of proteomic data in obtaining a holistic understanding of the molecular basis for tumor tissue morphology. Furthermore, survival analysis yielded specific morphological features related to patient prognosis, which have a strong association with important molecular events based on our analysis. Overall, our study demonstrated the power for integrating multiple types of biological data for cancer samples in generating new hypothesis as well as identifying potential biomarkers predicting patient outcome. Future work includes causal analysis to identify key regulators for cancer tissue development and validating the findings using more independent data sets.

Heterogeneous Effects of Fibroblast-Myocyte Coupling in Different Regions of the Human Atria Under Conditions of Atrial Fibrillation.

Background: Atrial fibrillation (AF), the most common cardiac arrhythmia, is characterized by alteration of the action potential (AP) propagation. Under persistent AF, myocytes undergo electrophysiological and structural remodeling, which involves fibroblast proliferation and differentiation, modifying the substrate for AP propagation. The aim of this study was to analyze the effects on the AP of fibroblast-myocyte coupling during AF and its propagation in different regions of the atria.Methods: Isolated myocytes were coupled to different numbers of fibroblasts using the established AP models and tissue simulations were performed by randomly distributing fibroblasts. Fibroblast formulations were updated to match recent experimental data. Major ion current conductances of the myocyte model were modified to simulate AP heterogeneity in four different atrial regions (right atrium posterior wall, crista terminalis, left atrium posterior wall, and pulmonary vein) according to experimental and computational studies.Results: The results of the coupled myocyte-fibroblast simulations suggest that a more depolarized membrane potential and higher fibroblast membrane capacitance have a greater impact on AP duration and myocyte maximum depolarization velocity. The number of coupled fibroblasts and the stimulation frequency are determining factors in altering myocyte AP. Strand simulations show that conduction velocity tends to homogenize in all regions, while the left atrium is more likely to be affected by fibroblast and AP propagation block is more likely to occur. The pulmonary vein is the most affected region, even at low fibroblast densities. In 2D sheets with randomly placed fibroblasts, wavebreaks are observed in the low density (10%) central fibrotic zone and when fibroblast density increases (40%) propagation in the fibrotic region is practically blocked. At densities of 10 and 20% the width of the vulnerable window increases with respect to control but is decreased at 40%.Conclusion: Myocyte-fibroblast coupling characteristics heterogeneously affect AP propagation and features in the different atrial zones, and myocytes from the left atria are more sensitive to fibroblast coupling.

Engineering the Interface: Nanodiamond Coating on 3D-Printed Titanium Promotes Mammalian Cell Growth and Inhibits Staphylococcus aureus Colonization.

Additively manufactured selective laser melted titanium (SLM-Ti) opens the possibility of tailored medical implants for patients. Despite orthopedic implant advancements, significant problems remain with regard to suboptimal osseointegration at the interface between the implant and the surrounding tissue. Here, we show that applying a nanodiamond (ND) coating onto SLM-Ti scaffolds provides an improved surface for mammalian cell growth while inhibiting colonization of Staphylococcus aureus bacteria. Owing to the simplicity of our methodology, the approach is suitable for coating SLM-Ti geometries. The ND coating achieved 32 and 29% increases in cell density of human dermal fibroblasts and osteoblasts, respectively, after 3 days of incubation compared with the uncoated SLM-Ti substratum. This increase in cell density complements an 88% reduction in S. aureus detected on the ND-coated SLM-Ti substrata. This study paves a way to create facile antifouling SLM-Ti structures for biomedical implants.

Role of the Notch Signaling Pathway in Fibrosis of Denervated Skeletal Muscle.

In order to investigate the role of the Notch signaling pathway in skeletal muscle fibrosis after nerve injury, 60 Sprague-Dawley rats were selected and divided randomly into a control and two experimental groups. Group A served as controls without any treatment. Rats in groups B were injected intraperitoneally with 0.2 mL PBS and those in group C were injected intraperitoneally with 0.2 mL PBS+100 μmol/L, 0.2 mL N-[N-(3,5-difluorophenacetyl)-l-alanyl]- S-phenylglycine t-butyl ester (DAPT, a gamma-secretase inhibitor that suppresses Notch signaling) respectively, on postoperative days 1, 3, 7, 10, and 14 in a model of denervation-induced skeletal muscle fibrosis by right sciatic nerve transection. Five rats from each group were euthanized on postoperative days 1, 7, 14, and 28 to collect the right gastrocnemii, and hematoxylin and eosin (HE) staining, immunohistochemistry test, real-time PCR, and Western blotting were performed to assess connective tissue hyperplasia and fibroblast density as well as expression of Notch 1, Jagged 1, and Notch downstream molecules Hes 1 and collagen I (COL I) on day 28. There was no significant difference in HE-stained fibroblast density between group B and C on postoperative day 1. However, fibroblast density was significantly higher in group B than in group C on postoperative days 7, 14, and 28. Notch 1, Jagged 1, Hes 1, and COL I proteins in the gastrocnemius were expressed at very low levels in group A but at high levels in group B. Expression levels of these proteins were significantly lower in group C than in group B (P<0.05), but they were higher in group C than in group A (P<0.05) on postoperative day 28. We are led to conclude that locking the Notch signaling pathway inhibits fibrosis progression of denervated skeletal muscle. Thus, it may be a new approach for treatment of fibrosis of denervated skeletal muscle.

Dielectric and Double Debye Parameters of Artificial Normal Skin and Melanoma

The aim of this study is to characterise the artificial normal skin and melanoma by testing samples with different fibroblast and metastatic melanoma cell densities using terahertz (THz) time-domain spectroscopy (TDS) attenuated total reflection (ATR) technique. Results show that melanoma samples have higher refractive index and absorption coefficient than artificial normal skin with the same fibroblast density in the frequency range between 0.4 and 1.6 THz, and this contrast increases with frequency. It is primarily because that the melanoma samples have higher water content than artificial normal skin, and the main reason to melanoma containing more water is that tumour cells degrade the contraction of the collagen lattice. In addition, complex refractive index and permittivity of the melanoma samples have larger variations than that of normal skin samples. For example, the refractive index of artificial normal skin at 0.5 THz increases 4.3% while that of melanoma samples increases 8.7% when the cell density rises from 0.1 to 1 M/ml. It indicates that cellular response of fibroblast and melanoma cells to THz radiation is significantly different. Furthermore, the extracted double Debye (DD) model parameters demonstrate that the static permittivity at low frequency and slow relaxation time can be reliable classifiers to differentiate melanoma from healthy skin regardless of the cell density. This study helps understand the complex response of skin tissues to THz radiation and the origin of the contrast between normal skin and cancerous tissues.

Effect of Honey/PVA Hydrogel Loaded by Erythromycin on Full-Thickness Skin Wound Healing in Rats; Stereological Study.

Background: Skin wounds are a significant public health risk, and treatment of wound remains a challenging clinical problem for medical teams and researchers. Materials and Methods: In the present study, we aimed to investigate the healing effects of honey/polyvinyl alcohol (PVA) hydrogel loaded with erythromycin as wound dressing on skin wounds in rats, based on histological studies. In this study, 60 male Wistar rats, with a 1.5 ×1.5 cm2 diameter full-thickness wounds on the backs were divided into four groups: honey/PVA with the erythromycin hydrogel group, honey group, PVA group, and the control group, with no treatment. Skin biopsies were prepared at days 4, 7, and 14 for microscopic analyses. The stereological analysis, including the mean area of the wound, length of vessels, numerical density of fibroblast, macrophage, basal cell and volume of the epidermis, dermis, and fibrous tissue were performed. Results: Wounds area in the honey/PVA hydrogel with the erythromycin group were significantly (P<0.05) smaller than in the other group. The numerical density of fibroblast, macrophage, basal cell and volume of the epidermis in the honey/PVA hydrogel with the erythromycin group were significantly higher than other groups. Conclusion: According to our results, honey/PVA hydrogel with erythromycin may promote early wound healing and has a positive influence on fibroblast proliferation and re-epithelialization, and its administration is recommended after further validation of clinical data.

Effect of Honey/PVA Hydrogel Loaded by Erythromycin on Full-Thickness

Background: Skin wounds are a significant public health risk, and treatment of wound remains a challenging clinical problem for medical teams and researchers.Materials and Methods: In the present study, we aimed to investigate the healing effects of honey/polyvinyl alcohol (PVA) hydrogel loaded with erythromycin as wound dressing on skin wounds in rats, based on histological studies. In this study, 60 male Wistar rats, with a 1.5 ×1.5 cm2 diameter full-thickness wounds on the backs were divided into four groups: honey/PVA with the erythromycin hydrogel group, honey group, PVA group, and the control group, with no treatment. Skin biopsies were prepared at days 4, 7, and 14 for microscopic analyses. The stereological analysis, including the mean area of the wound, length of vessels, numerical density of fibroblast, macrophage, basal cell and volume of the epidermis, dermis, and fibrous tissue were performed. Result: Wounds area in the honey/PVA hydrogel with the erythromycin group were significantly (P&lt;0.05) smaller than in the other group. The numerical density of fibroblast, macrophage, basal cell and volume of the epidermis in the honey/PVA hydrogel with the erythromycin group were significantly higher than other groups.Conclusion: According to our results, honey/PVA hydrogel with erythromycin may promote early wound healing and has a positive influence on fibroblast proliferation and re-epithelialization, and its administration is recommended after further validation of clinical data.[GMJ. 2019;8:e1362]

Matricaria Chamomilla Extract Improves Diabetic Wound Healing in Rat Models

Background: Diabetes mellitus (DM) has a broad spectrum of complications and comorbidities. Delayed wound healing is a major reason for infections and further life-threatening complications; the process of vascularization and tissue regeneration as well as anti-inflammatory and anti-oxidant reactions of the body are impaired in DM. Matricaria Chamomilla (MC) is a herbal medicine which has antimicrobial, antioxidant and wound healing potentials.Objectives: In this study, these potentials are evaluated on full thickness diabetic excisional skin wounds by using stereological methods.Methods: In this experimental study, 48 male Wistar rats with streptozotocin (STZ)-induced diabetes (50 mg/kg intraperitoneally) were divided into 4 groups (n = 12) randomly: MC 5% gel treated, MC 10% gel treated, gel-base treated group, and the control group receiving no treatment. After creating 1 cm2 wounds, rats were treated every 24 hours for 15 days. Volume densities of collagens, wound closure rate, volume density, length density, and mean diameter of vessels, and fibroblast proliferation, were examined and analyzed. P ≤ 0.05 was mentioned as statistically significant.Results: MC gels receiving groups showed hastened wound area reduction in contrast with gel-base and control groups (P < 0.05). Numerical density of fibroblasts, mean diameter and length densities of the vessels in MC groups were significantly higher than the control and the base groups (P < 0.05).Conclusions: It is demonstrated that MC has the ability to improve wound healing by enhancing fibroblast proliferation and revascularization in diabetic skin injuries and has the potential to be considered as an alternative treatment.

Ultrastructural and histological changes in tibial remnant of ruptured anterior cruciate ligament stumps: a transmission electron microscopy and immunochemistry-based observational study.

Anterior cruciate ligament (ACL) rupture is a common injury and has a non-union rate of 40-100%. Important cellular events, such as fibroblast proliferation, angiogenesis and change in collagen fibril thickness in the ACL remnant, as described in other dense connective tissue, might have an implication in graft recovery following ACL reconstruction. Thus we conducted a study with an aim to characterize the ultrastructural and histological features of ruptured ACL tibial stump and correlate the same with the duration of injury. This was a prospective observational study in which 60 ruptured human ACLs were evaluated for collagen fibril thickness, blood vessel density (per mm2) and fibroblast density (per mm2) with the help of transmission electron microscopy, immunohistochemistry via CD34 antibody staining and light microscopy (H&E staining). The findings were correlated with duration of injury. Fifty-four male and six female patients with a mean duration of the injury of 23.01weeks (SD = 26.09; range 2-108weeks) were included for the study and were divided on the basis of duration of injury as follows: Group I (≤6weeks; N = 16), Group II (7-12weeks; N = 18), Group III (13-20weeks; N = 7), Group IV (21-50weeks; N = 12), Group V (> 50weeks; N = 7). A significant correlation was seen with blood vessel density (r = 0.303, p = 0.01) and fibroblast density (r = - 0.503, p = 0.001). Thickness of collagen fibril did not correlate with the duration of injury (r = 0.15, p = 0.23). The thickness of the collagen reached its peak after 50weeks following injury, whereas highest density of blood vessel and fibroblast was seen at 12-20weeks. Matched pair analysis revealed a significant decrease in collagen fibril thickness and an increase in fibroblast density at 7-12weeks. Following injury to ACL, the ruptured tibial stump undergoes a series of changes at the cellular level vis-à-vis changes in collagen fibril thickness, vascular density and fibroblast density that possibly suggest an intrinsic healing response. This further may have implications on the functional outcome following ACL reconstruction with remnant preservation. III.

Effect of different plasma treated zirconia on the adhensive behaviour of human gingival fibroblasts

OBJECTIVE To evaluate the effect of different cold atmospheric plasma (CAP) treatment on the surface chemical and physical properties of zirconia and adhensive behaviour of human gingival fibroblasts (HGFs) cultured on zirconia disks. METHODS The zirconia disks were divided into four groups and treated using helium, argon and mixture of argon and oxygen cold atmospheric plasma for 90 s or left untreated. The surface morphology, wettability and chemical elements were analyzed right after treatment. Human gingival fibroblasts were grown from biopsies obtained from a periodontally healthy human subject during periodontal surgery. HGFs were seeded on zirconia disk, and cells density was measured at the time point of 3 hours. Indirect immunofluorescence (IIF) was performed for morphometric examination at the time point of 3 hours. RESULTS The crystallographic structure of zirconia was analyzed previously and the results suggested that it fitted the properties of zirconium yttrium oxide. After helium, argon and mixture of argon and oxygen cold atmospheric plasma treatment, the surface morphology and roughness of zirconia disks remained the same. The contact angle of zirconia decreased significantly(P<0.05)after CAP treatment: from 68.38° to 17.90°. After different CAP plasmas treatment, the atomic percentage of carbon on the outermost surface of the three groups decreased, as did the surface C/O ratio. The surface C/O ratio of zirconia decreased from 1.07 to 0.33. Fibroblasts density increased on CAP treated disks, especially the ones treated by mixture of argon and oxygen CAP(P<0.05). Cells of the three CAP plasma treatment groups spread better and had more protrusions, as well as larger surficial areas. CONCLUSION Based on the results of this study after being treated by different kinds of CAP plasmas for 90 s, the surface wettability increased and the elements changed significantly with no changes in the tomography and roughness of the materials. The CAP treatment enhances the adhensive behavior of fibroblasts on zirconia by increasing the oxygen functional groups and promoting the cell density. Wettability of zirconia, an important physicochemical property, has a vital influence on the cell behaviors.

The role of beluntas (pluchea indica less.) leaf extract in preventing the occurrence of fibroblasts hyperproliferation: An in vitro preliminary study

Beluntas (Pluchea indica Less.) is a herbal plant which contains variety of benefits. Quercetin, one of flavonoid, is the most bioactive agent in beluntas leaf. Collagen inhibition by quercetin may modulate extracellular matrix deposition and inhibit the formation of hypertrophic scar. This was an in vitro study with senescent fibroblasts to determine the role of beluntas leaf extract in preventing the occurence of fibroblasts hyperproliferations. There were 4 groups were stained by anti-collagen I antibodies and secondary antibody. Flowcytometry analysis was done to measure the fibroblasts density. Anova test was performed with a value of p=0.000 (p&lt;0.05). A post hoc analysis showed significant differences in the average decrease of fibroblasts that absorbs staining anti-collagen I antibody treatment group compared with the control group. There were significant effects of beluntas leaf extract in preventing the occurrence of fibroblasts hyperproliferations. Beluntas leaf extract with a concentration of 80 mol/L had the most significant effect on the fibroblasts density. Thus beluntas leaf extract has the ability in preventing the occurrence of fibroblasts hyperproliferation.

Self-organization of conducting pathways explains electrical wave propagation in cardiac tissues with high fraction of non-conducting cells

Cardiac fibrosis occurs in many forms of heart disease and is considered to be one of the main arrhythmogenic factors. Regions with a high density of fibroblasts are likely to cause blocks of wave propagation that give rise to dangerous cardiac arrhythmias. Therefore, studies of the wave propagation through these regions are very important, yet the precise mechanisms leading to arrhythmia formation in fibrotic cardiac tissue remain poorly understood. Particularly, it is not clear how wave propagation is organized at the cellular level, as experiments show that the regions with a high percentage of fibroblasts (65-75%) are still conducting electrical signals, whereas geometric analysis of randomly distributed conducting and non-conducting cells predicts connectivity loss at 40% at the most (percolation threshold). To address this question, we used a joint in vitro-in silico approach, which combined experiments in neonatal rat cardiac monolayers with morphological and electrophysiological computer simulations. We have shown that the main reason for sustainable wave propagation in highly fibrotic samples is the formation of a branching network of cardiomyocytes. We have successfully reproduced the morphology of conductive pathways in computer modelling, assuming that cardiomyocytes align their cytoskeletons to fuse into cardiac syncytium. The electrophysiological properties of the monolayers, such as conduction velocity, conduction blocks and wave fractionation, were reproduced as well. In a virtual cardiac tissue, we have also examined the wave propagation at the subcellular level, detected wavebreaks formation and its relation to the structure of fibrosis and, thus, analysed the processes leading to the onset of arrhythmias.

TNF-α-Activated MSC-CM Topical Gel Effective in Increasing PDGF Level, Fibroblast Density, and Wound Healing Process Compared to Subcutaneous Injection Combination

Mesenchymal stem cells (MSCs) are multipotent stromal cells that have the capacity to regenerate tissue damage. However, they have several limitations. MSC-CM as a new approach treatment is widely used to solve the limitation of MSC in wound healing. The aim of this study was to evaluate the effectiveness of TNF-α-activated MSC-CM topical gel compare to topical-subcutaneous injection combination on wound healing acceleration. This study was conducted between April and August 2018 at the Stem Cell and Cancer Research Laboratory (SCCR), Faculty of Medicine, Sultan Agung Islamic University, Semarang. Experimental post-test only control group design was performed by involving 36 animal models randomly divided into six groups; T1, T2 (MSC-CM in topical gel 100 μL; 200 μL); ST1, ST2 (MSC-CM in subcutaneous injection : topical gel = 80 μL:20 μL; 160 μL:40 μL); CT (200 μL medium free TNF-α); CST (PBS in subcutaneous injection : topical gel = 160 μL :40 μL). The measurement of PDGF level on day 3 and 6 was conducted using ELISA assay while the fibroblast density was analyzed by light microcopy. It was found that there was was a significant increase in PDGF and fibroblast density on day 6 in the topical group when compared to the combination group (p<0,05). It is concluded that the MSC-CM topical gel is more effective than combination of topical-subcutaneous injection. Key words: Combination, fibroblast, MSC-CM, PDGF, subcutaneous MSC-CM, topical MSC-CM MSC-CM Topikal yang diaktivasi TNF-α Efektif Dalam Peningkatan Level PDGF, Densitas Fibroblast, dan Mempercepat Penyembuhan Luka dibanding dengan Kombinasi Injeksi Subkutan Mesenchymal stem cells (MSCs) merupakan sel stroma multipoten yang memiliki kemampuan untuk meregenerasi kerusakan jaringan. Namun, MSC memiliki beberapa keterbatasan. MSC-CM sebagai terapi pendekatan baru digunakan untuk mengatasi keterbatasan MSC dalam penyembuhan luka. Tujuan penelitian ini adalah menilai efektivitas MSC-CM topikal yang diaktvasi TNF-α dibandingdengan kombinasi topikal-injeksi subkutan pada percepatan penyembuhan luka. Penelitian ini dilaksanakan pada bulan April–Agustus 2018 di Laboratorium Stem Cell and Cancer Research (SCCR), Fakultas Kedokteran, Universitas Islam Sultan Agung, Semarang. Penelitian menggunakan eksperimen laboratorium dengan rancangan post-test only control group, menggunakan 36 tikus galur wistar yang dibagi secara acak menjadi 6 kelompok; T1, T2 (MSC-CM gel topikal 100μL; 200μL); ST1, ST2 (MSC-CM injeksi subkutan : gel topikal = 80 μL:20 μL; 160 μL:40 μL); CT (200 μL medium tanpa TNF-α); CST (PBS injeksi subkutan: gel topikal = 160 μL :40 μL). Pengukuran kadar PDGF pada hari ke-3 dan ke-6 mengunakan ELISA, sedangkan jumlah fibroblas dilihat mengunakan mikroskop cahaya. Hasil penelitian ini menunjukkan peningkatan kadar PDGF dan jumlah fibroblas yang signifikan dihari ke-6 pada MSC-CM gel topikal dibanding dengan kombinasi topical-injeksi subukutan (p <0.05). Simpulan penelitian ini adalah pemberian MSC-CM secara topical lebih efektif dibanding dengan kombinasi topikal-injeksi subkutan. Kata kunci: Fibroblas, konditional medium, MSC-CM kombinasi, MSC-CM topikal, PDGF