Thick Collagen Fibers Research Articles

Effect of Vitamin D Supplementation on Doxorubicin-Induced Cardiotoxicity in Rats

Abstract Background Doxorubicin-induced cardiotoxicity is a worldwide problem. Vitamin D is a well-known beneficial vitamin for bone growth and calcium homeostasis but recently it is also known for its cardioprotective effects. The aim of this study is to investigate the potential protective role of vitamin D on the cardiac dysfunction induced by chronic doxorubicin exposure, and to throw more light on the possible underlying mechanism (s) for such effect. Materials and Methods: 70 female Albino-rats were divided into 4 groups; control group (C), Doxorubicin-treated group (Dox): given i.p. injection of Dox in a dose of 2.5 mg/kg body weight (cumulative dose: 15 mg/kg) over 3 weeks, vitamin Dsupplemented group (Vit D): given vitamin D by oral gavage in a dose of 500 IU/kg daily, 5 days a week, also for 3 weeks and the combined Doxorubicintreated+vitamin D-supplemented group (Dox+Vit D). At the end of the experiment, ECG was recorded and in vitro isolated heart study was performed on Langendoroff preparation to measure peak tension (PT), time to peak tension (TPT), half relaxation time (HRT) and myocardial flow rate (MFR). Body and cardiac weights, plasma levels of brain naturetic peptide (BNP), cardiac troponin I (cTnI), vitamin D and total calcium and cardiac tissue heat shock protein 20, total antioxidant capacity (TAC) and malondialdehyde (MDA) were measured. Also, cardiac tissues were histopathologically assessed. Results: Dox-treated rats showed significant decrease in the final body weight (fBW), significant prolongation of the P-R interval, QRS duration, observed Q-T (Q-TO) and corrected Q-T (Q-Tc) with significant depression of the R voltage. In addition, there was a significant decrease in the in vitro heart rate, significant depression in PT, PT/LV and MFR together with significant prolongation in TPT& 3 HRT. These changes were accompanied by significant elevation of plasma BNP, cTnI and in cardiac tissue MDA and a significant decrease in plasma vit D, total calcium and cardiac tissue TAC and HSP20. Histopathological examination revealed markedly distorted muscle fibers with indistinct cell borders, bright eosinophilic cytoplasm, intra-cytoplasmic vacuoles and small pyknotic nuclei or absent nuclei, together with interstitial edema & aggregates of inflammatory cells and thick irregular collagen fibers in between the muscle fibers. Concomitant supplementation of vitamin D to the doxorubicin treated rats resulted in significant decrease in PR interval, QRS duration, MDA and significant increase PT, PT/LV, MFR, MFR/LV, plasma vitamin D, total calcium and TAC compared to the Dox treated rats to be insignificantly different from the control group. Plasma BNP and cTnI were significantly decreased while cardiac HSP20 was significantly increased compared to the Dox-treated rats, yet these parameters were still significant from the control group. Meanwhile, fBW, Q-TO and Q-Tc intervals, and TPT remained insignificantly changed from the DOX group. These findings were associated by regaining the normal collagen fiber distribution between cardiac muscle fibers with resolution of interstitial edema. Conclusion: Vitamin D supplementation can partially mitigate cardiac dysfunction induced by chronic doxorubicin by improving the cardiac antioxidant state and heat shock protein 20 level. Key words: Doxorubicin, cardiac dysfunction, vitamin D, isolated heart studies, BNP, HSP20.

Intrauterine exposure to chronic hypoxia in the rat leads to progressive diastolic function and increased aortic stiffness from early postnatal developmental stages.

AimWe sought to explore whether fetal hypoxia exposure, an insult of placental insufficiency, is associated with left ventricular dysfunction and increased aortic stiffness at early postnatal ages.MethodsPregnant Sprague Dawley rats were exposed to hypoxic conditions (11.5% FiO2) from embryonic day E15‐21 or normoxic conditions (controls). After delivery, left ventricular function and aortic pulse wave velocity (measure of aortic stiffness) were assessed longitudinally by echocardiography from day 1 through week 8. A mixed ANOVA with repeated measures was performed to compare findings between groups across time. Myocardial hematoxylin and eosin and picro‐sirius staining were performed to evaluate myocyte nuclear shape and collagen fiber characteristics, respectively.ResultsSystolic function parameters transiently increased following hypoxia exposure primarily at week 2 (p < .008). In contrast, diastolic dysfunction progressed following fetal hypoxia exposure beginning weeks 1–2 with lower early inflow Doppler velocities, and less of an increase in early to late inflow velocity ratios and annular and septal E’/A’ tissue velocities compared to controls (p < .008). As further evidence of altered diastolic function, isovolumetric relaxation time was significantly shorter relative to the cardiac cycle following hypoxia exposure from week 1 onward (p < .008). Aortic stiffness was greater following hypoxia from day 1 through week 8 (p < .008, except week 4). Hypoxia exposure was also associated with altered nuclear shape at week 2 and increased collagen fiber thickness at week 4.ConclusionChronic fetal hypoxia is associated with progressive LV diastolic dysfunction, which corresponds with changes in nuclear shape and collagen fiber thickness, and increased aortic stiffness from early postnatal stages.

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.&#x0D; 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.&#x0D; 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).&#x0D; 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.&#x0D; 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.&#x0D; 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.&#x0D; Conclusions&#x0D; &#x0D; 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.&#x0D; 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.&#x0D; 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.&#x0D;

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.&#x0D; 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.&#x0D; 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).&#x0D; 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.&#x0D; 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.&#x0D; 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.&#x0D; Conclusions&#x0D; &#x0D; 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.&#x0D; 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.&#x0D; 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.&#x0D;

2010P - Quantitative imaging and characterization of collagen patterns in high grade serous ovarian carcinoma (HGSOC)

BackgroundHGSOC is known to demonstrate diverse molecular heterogeneity. It is unclear whether the tumor microenvironments such as the stromal components in the extracellular matrix (ECM) also exist heterogeneity. MethodsAs a proof-of concept study, characterization of collagen patterns was performed on 60 unstained formalin-fixed paraffin embedded (FFPE) HGSOC samples (three sections from each of the 20 patients). Each section with 5 micron thickness and a minimum tumour surface area of 40mm2 were scanned by using a multiphoton imaging system (Genesis 200, HistoIndex) following deparaffinization. Parameters including Collagen Area Ratio (CAR), Collagen Fiber Density (CFD), Collagen Reticulation Index (CRI), Collagen Fiber Number (CFN), Collagen Fiber Thickness (CFT), and Collagen Fiber Length (CFL) were analysed. Unsupervised hierarchical clustering analysis was performed. ResultsUnsupervised hierarchical clustering of the collagen parameters revealed four distinct patterns in HGSOC samples. G1 tumors consisted of long and thick collagen fibers with high CFT and CFL. G2 tumors were low in all the collagen parameters, suggesting a relatively “clean” stromal without collagen deposition. G3 tumors consisted of dense collagen fibers with high CRI suggestive of extensive cross alignment among the fibers. G4 tumors were high in CFN and low in CRI, CFT and CFL, suggesting a stroma loaded with high amount of thin and short collagen fibers without cross alignment.The collagen patterns were not exclusive for specific organ sites (ovary, fallopian tube, other metastatic sites) except that the collagen pattern from the omental metastases were mainly G2 (10/18; 55.6%) and G4 (6/18; 33.3%). ConclusionsThe stromal component in the ECM of HGSOC can be successfully quantitated by the multi-photon imaging technology on unstained FFPE sections. The collagen component exhibited significant heterogeneity in terms of the number, thickness, length, and reticulation patterns. The contribution of different collagen patterns to clinical outcomes and the correlation with known molecular subtypes in HGSOC warrants further investigation in larger cohorts. Legal entity responsible for the studyNational University of Singapore. FundingNational Medical Research Council of Singapore. DisclosureAll authors have declared no conflicts of interest.

Inhibition of transglutaminase activity in periodontitis rescues periodontal ligament collagen content and architecture.

Periodontal disease (PD) afflicts approximately 50% of the population in the United States and is characterized by chronic inflammation of the periodontium that can lead to loss of the periodontal ligament through collagen degradation, loss of alveolar bone, and to eventual tooth loss. Previous studies have implicated transglutaminase (TG) activity in promoting thin collagen I fiber morphology and decreased mechanical strength in homeostatic PDL. The aim of this study was to determine whether TG activity influenced collagen assembly in PDL in the setting of periodontal disease. A ligature model was used to induce clinically relevant PD in mice. Mice with ligature were assessed at 5 and 14days to determine PDL collagen morphology, transglutaminase (TG) activity, and bone loss. The effects of inhibition of TG on PDL were assessed by immunohistochemistry and second-harmonic generation (SHG) to visualize collagen fibers in native tissue. Ligature placement around the 2nd molar resulted in significant bone loss and a decrease in total collagen content after 5days of ligature placement. A significant increase in thin over thick fibers was also demonstrated in mice with ligature at 5days associated with apparent increases in immunoreactivity for TG2 and for TG-mediated N-ε-γ-glutamyl cross-links in PDL. Inhibition of TG activity increased total collagen and thick collagen fiber content over vehicle control in mice with ligature for 5days. SHG of PDL was used to visualize and quantify the effects of TG inhibition on enhanced collagen fiber organization in unfixed control and diseased PDL. These studies support a role of TG in regulating collagen fiber assembly and suggest that strategies to inhibit TG activity in disease might contribute to restoration of PDL tissue integrity.

Superficial Morphea: Clinicopathological Characteristics and a Novel Therapeutic Outcome to Excimer Light Therapy.

Introduction Superficial morphea (SM) is an uncommon entity that was described in the literature without definitive correlation to localized scleroderma (LS) or other atrophoderma diseases. Aim To demonstrate the clinicopathological features of SM and evaluate the efficacy of different therapeutic modalities in its management. Patients and methods A total of 28 patients with SM were studied during the period from 2010 to 2015. Clinicopathological features and therapeutic outcomes were recorded and analyzed. Results Clinically, SM was predominant in females (71.4%) with an average onset at 33 years of age and an average duration of 15 months. It was commonly presented as asymptomatic, darkly pigmented, and multiple and slightly indurated patches. The lesions were mostly ill-defined, large-sized, and located more on the trunk. Histologically, thickening of collagen fibers was observed either localized to the papillary dermis only (38.9%) or extended into the upper reticular dermis (61.1%). Elastic fibers were generally diminished in the upper reticular dermis while the number of fibroblasts and basal melanin pigmentation were increased in the majority of cases (92.9% and 96.4%, respectively). The most commonly associated diseases were diabetes mellitus (50%) and hepatitis C virus (HCV) infection (42.8%), and their incidence was significantly higher than that in patients with LS. Excimer light showed promising effective results in the treatment of most cases (78.9%) while the response to other modalities such as topical corticosteroid alone or in combination with tacrolimus or treatment with UVA1 alone was less effective (7.1%, 23.1%, and 5%, respectively). Conclusion Our results proposed that SM is a distinctive clinicopathological variant and not a stage in the spectrum of LS. The novel response of SM to excimer light and not for UVA1 therapy also suggests the different therapeutic outcome of SM from LS. Although SM has a significant association with DM and HCV infection, they seem not to affect the course of the disease.

Topical hydrogel containing Fumaria vaillantii Loisel. extract enhances wound healing in rats

BackgroundFumaria species (Fumariacea) has traditionally been used in wound healing in Iranian folk medicine. However, with the discovery of newer agents, its use has faded off into total obscurity. This study explored the wound healing potential of a gel containing 10% Fumaria vaillantii Loisel through topical application of total extract in a model of excisional as well as incisional wound healing in albino Wistar rats.MethodsRats were anesthetized, and excisional skin wound was established using a sterilized surgical scissors. The animals were then treated with 10% F.vaillantii topical gel formulation along with the gel base. The treatments were administered once a day after the injury for 21 days. For topical treatment, the hydrogel was formulated and evaluated for chemical and physical characteristics. Histopathological analysis with hematoxylin and eosin (H&E) was used for microscopic examination of the skin tissues on 21-day-old sections of excision wound. To verify collagen formation, hydroxyproline determination was performed 21 days post wound healing. Breaking strength was determined in a 10-day-old incision wound by the uniaxial tensile test.ResultsTopical administration of F.vaillantii gel formulation significantly enhanced skin wound closure on the 6th post-wounding day compared to both gel base and the negative control, indicating an accelerated wound healing process, while a significant difference was observed on 10th and 14th post –wound days in F.vaillantii treatment compared to the negative control groups. Gel formulation prepared with a 10% F. vaillantii extract exhibited a response in terms of wound epithelialization, angiogenesis and number of hair follicles at wound area better than the gel base on the 21st post-wound day. Application of gel base produced further advantages by increasing hydroxyproline content and collagen fiber thickness. Our results on incision wound model were supported by histopathological data indicating the role of gel base in the enhancement of breaking strength.ConclusionTraditional use of Fumaria species in the skin diseases was justified in this study by revealing the increase in wound healing activity after hydrogel containing F. vaillantii total extract administration.Graphical abstract

Characterizing differences in the collagen fiber organization of skin wounds using quantitative polarized light imaging.

Collagen fiber organization requires characterization in many biomedical applications, but it is difficult to objectively quantify in standard histology tissue sections. Quantitative polarized light imaging is a low-cost technique that allows for rapid measurement of collagen fiber orientation and thickness. In this study, we utilize a quantitative polarized light imaging system to characterize fiber orientation and thickness from wound sections. Full thickness skin wound sections that were previously stained with hematoxylin and eosin were used to assess collagen fiber content and organization at different points during the wound healing process. Overall, wounds exhibited a measurable increase in collagen fiber thickness and a nonlinear change in fiber reorganization within the wound. Our study demonstrates that quantitative polarized light imaging is an inexpensive alternative or supplement to standard histology protocols, requiring no additional stains or dyes, and yields repeatable quantitative assessments of collagen organization.

Abstract P157: Role of P47 phox in Angiotensin II Induced Cardiac Hypertrophy and Fibrosis in Mice

NADPH oxidase mediated ROS production has been recognized to be an important contributory mechanism for hypertension and left ventricular hypertrophy (LVH). However, the regulatory role of NADPH oxidase mediated superoxide production in Angiotensin II (AngII) induced cardiac fibrosis and macrophage infiltration is still elusive. In the present study, we investigated the role of phagocytic NADPH oxidase enzyme system in cardiac remodelling induced by AngII in young adult (2-3 months old) male mice deficient in p47 phox , a cytosolic subunit of the NADPH oxidase (KO, n=13) and age-matched wild-type littermates (WT, n=9). Animals were infused with AngII pressor dose (1500 ng/kg/min, s.c) or saline via osmotic minipump (Alzet 2002, USA) for 14 consecutive days and then, subjected to echocardiography (VEVO 1100, Canada). Further animals were sacrificed and hearts was harvested for histological studies and gene expression analysis. AngII treatment in WT and KO mice showed prominent alterations in echocardiographic measurements with no changes in E/A, EF and FS. However, KO mice showed aggravated cardiac hypertrophic response (LVAWd: WT, 1.20±0.09 vs. KO, 1.42±0.04, p&lt;0.05; LVIDd: WT, 3.02±0.20 vs. KO, 2.60±0.07, p&lt;0.05; LVPWd: WT, 1.49±0.06 vs. KO, 1.78±0.06, p&lt;0.05). Histologically, AngII infused KO mice also showed increased cardiomyocyte diameter compared to AngII infused WT mice (WT, 388±16.37 vs. KO, 560.3±41.28 μm 2 ; p&lt;0.01 respectively). Further, AngII infused KO mice showed elevated immune cell infiltration, which were positive for Mac-3 staining compared to AngII infused WT mice (WT, 2.61±0.32 vs. KO, 4.09±0.44 %; p&lt;0.05). Besides, AngII infused KO mice showed augmented interstitial fibrosis (WT, 11.11±0 0.67 vs. KO, 20.85± 3.63 %; p&lt;0.05), thick collagen fibres deposition (WT, 28.93± 0.94 vs. KO, 37.60± 2.49 %; p&lt;0.05) compared to AngII infused WT mice. Moreover, AngII infused KO mice also showed upregulated gene expression of hypertrophic and fibrotic markers, including Nppa, Nppb, Acta1, Myh6, TGF-β1, collagen (I and III) and α-sma compared to AngII infused WT mice. Together, these data suggest that NADPH oxidase and its subunit p47 phox play a pivotal role in AngII induced LVH by regulating fibrotic machinery and macrophage infiltration.

Lamina propria: The connective tissue of rat urinary bladder mucosa.

To describe and illustrate the structure of the propria, the bladder of adult rats was fixed in controlled conditions of distension and examined by light and electron microscopy. The lamina propria, ~50 µm thick in the distended bladder, consists of a superficial part (the cellular component), adjacent to the urothelium, rich in nerves, capillaries, fibroblasts and thin bundles of collagen, and a deep, thicker part (the fibrous component), adjacent to the detrusor, rich in large collagen fibres and with few fibroblasts. In the cellular part there is an extensive plexus of afferent nerve fibers and a dense capillary network (with numerous pericytes), lying close to the urothelium, that is unique to the bladder. The main resident cells are fibroblasts, adhering to each other at the end of laminar extensions without forming specialized junctions. The deep part of the lamina propria is made of thick collagen fibers, interwoven and crisscrossing each other, with a few fibroblasts in the interstitial spaces between them. In summary, the superficial part of the lamina propria has most of the bladder afferent nerves, contains many fibroblasts and has a network of suburothelial capillaries. The deep part as a whole forms an ovoid balloon of woven fibrous material that is acted upon by the detrusor musculature attached to its outer surface. The lamina propria is a strong fibrous barrier between urothelium and musculature. The abundance of collagen points to the main role for its fibroblasts, that is, the production of collagen fibrils, assisting the mechanical role of the lamina propria.

Myocardial interstitial matrix as novel target for succinic acid treatment strategies during experimental hypobaric hypoxia

Objective — Quantitative morphological studies of myocardial cellular elements. Aim of research is to study myocardial interstitial matrix with and without succinic acid treatment in the rats exposed to intermittent hypobaric hypoxia. Material and Methods — The study was conducted on 26 adult males of Wistar rats weighing 220-310 g, divided into 3 groups. The first control group consisted of 6 intact animals. The second group included 10 rats which were exposed to hypobaric hypoxia without medication for 30 days. Third group was composed of 10 rats, which were medicated by succinic acid solution which was injected intraperitoneally once a day at the rate of 0.5 ml/100 g of animal body weight 15 minutes before hypoxic exposure for 30 days. Results — In the second group almost the entire stroma was consisting of the thickened collagen fibers and proliferating cells of the connective tissue. Their total number increased relatively to the control by 31% initiating the processes of collagenases. An increase in the number of myofibroblasts which was observed in the hypoxic rat heart in the second group followed by high MPP-9 expression. Hypoxic effects in the third group were not significantly different from control according to the observed changes in the stroma. In rats of the third group the capillary diameter exceeded the indicator of the second group, but was significantly lower than the control indicator. Accordingly, compared with the series without correction, the cross-sectional area of the capillaries was increased – 28.44±0.14 μm2 against 24.53±0.20 μm2. The total cross-sectional area of the capillaries is 0.77±0.10 x103 μm2. The relative surface area of the vascular area was lower than the control by 21.8%. Conclusion ― Fibrosis in the myocardium inevitably leads to increased myocardial stiffness, resulting in systolic and diastolic dysfunction, neurohormonal activation and, ultimately, heart failure caused by hypobaric hypoxia. Reduced oxygen delivery by microvascular damage, increased perivascular fibrosis may contribute to contractile failure. Succinic acid combined with inosine acts as a high-energy phosphate reserve, to maintain adenosine triphosphate at levels sufficient to support contractile function.

The structure and properties of natural sheep casing and artificial films prepared from natural collagen with various crosslinking treatments

The structure and properties of natural sheep casing and collagen films with various crosslinking treatments have been investigated in detail to develop satisfied artificial casings prepared from collagen. The sheep casing consists of large number of thick collagen fibers oriented at ±45° from longitudinal direction with high-density interwoven network structure. The structural feature of sheep casing gave the special mouthfeel of ‘cracking bite’ of sausages. Whereas, layered structure filled with fine collagen fibrils and large gaps in collagen film results in poor mechanical properties and higher swelling ratio in water. Furthermore, a degree of denaturation of collagen during extraction process also lead to poor mechanical properties. After glutaraldehyde (GTA) and dehydrothermal (DHT) treatments, the formation of crosslinking improved mechanical properties of collagen films significantly and the tensile strength and tensile modulus increased more than three times compared with those of untreated collagen film in wet before and after boiling. The swelling ratio of treated collagen films also decreased dramatically. No obvious effects on denaturation of collagen film after GTA treatment, but the degree of denaturation of DHT treated collagen film increased slightly.

Finasteride-Induced Inhibition of 5α-Reductase Type 2 Could Lead to Kidney Damage-Animal, Experimental Study.

In the pharmacological treatment of prostate cancer, benign prostatic hyperplasia and androgenetic alopecia finasteride is commonly used. This drug inhibits 5α-reductase type 2, which is why finasteride affects androgen homeostasis, since testosterone (T) cannot be reduced to dihydrotestosterone (DHT). As studies on sex-related renal injuries suggest a high probability of androgen-induced renal dysfunction, the aim of this study was to determine the potential harmful effects of finasteride on the kidneys of rats. The study was performed on sexually mature male Wistar rats given finasteride. Histological sections of the kidneys were used for immunohistochemical visualization of the androgen receptor (AR), junctional proteins (occluding (Occ); E-cad, N-cad, E-/N-cadherin; β-cat, β-catenin; connexin 43 (Cx43)), proliferating cell nuclear antigen (PCNA), IL-6, and lymphocyte markers (CD3 for T cell, CD19 for B cell). The TUNEL method was used for cell apoptosis identification, and picro sirius red staining was used to assess collagen fibers thickness. The levels of T, DHT and estradiol (E2) were determined in blood serum. It was shown that finasteride treatment affected steroid hormone homeostasis, altered the expression of AR and intracellular junction proteins, changed the ratio between cell apoptosis and proliferation, and caused lymphocyte infiltration and an increase of IL-6. The thickening of collagen fibers was observed as tubular fibrosis and glomerulosclerosis. Summarizing, finasteride-induced hormonal imbalance impaired the morphology (i.e., dysplastic glomeruli, swollen proximal convoluted tubules) and physiology (changed level of detected proteins/markers expression) of the kidneys. Therefore, it is suggested that patients with renal dysfunction or following renal transplantation, with androgen or antiandrogen supplementation, should be under special control and covered by extended diagnostics, because the adverse negative effect of DHT deficiency on the progression of kidney disease cannot be ignored.

Stromal Changes In Benign and Malignant Salivary Gland Tumors With Picrosirius Red-A Polarizing Microscopic Study

Objectives: To explore the polarizing color profile in connective tissue stroma of Picro-Sirius Red (PSR) stained sections of benign and malignant salivary gland tumors (SGT) and to assess the possible utility of this method as a diagnostic tool. Materials and Methods: A total number of 60 Paraffin blocks of benign (30 cases) and malignant (30 cases) salivary gland tumors were selected from the archives of our institute and regional cancer hospital. 30 cases of Mucous Extravasation Phenomenon (MEP) were selected as control. Salivary gland tumors and control were stained with PSR stain and examined under polarizing microscope. We counted 50 thin and 50 thick collagen fibers in each section of benign and malignant SGT and were categorized as Greenish Yellow (GY) or Yellowish Orange (YO). Statistical analysis was performed by using oneway ANOVA and Paired t-test. Results: We found similar presentation of polarizing color arrangement for thin collagen fibers (80-85% GY and 15-20% YO) in both benign and malignant SGT and MEP. In malignant SGT, thick collagen fibers exhibited different polarizing color (80% GY) than benign SGT and MEP (12-14% GY). Polarizing color profile of collagen in malignant SGT displayed a different presentation as compared to benign SGT and control (MEP) and differences were statistically significant(p<0.05). Conclusion: This study concluded about the difference of stromal collagen fibers in benign and malignant SGT. Malignant SGT suggesting loosely arranged collagen fibers could be involved in further growth and invasion of disease along with other cellular and molecular events. Keywords: Collagen, Mucous Extravasation Phenomenon, Picro-Sirius Red (PSR) Stain, Polarized Microscopy, Salivary Gland Tumor

Neuronal PAS Domain 2 (Npas2)-Deficient Fibroblasts Accelerate Skin Wound Healing and Dermal Collagen Reconstruction.

The circadian clock, which consists of endogenous self-sustained and cell-autonomous oscillations in mammalian cells, is known to regulate a wide range of peripheral tissues. The unique upregulation of a clock gene, neuronal PAS domain protein 2 (Npas2), observed along with fibroblast aging prompted us to investigate the role of Npas2 in the homeostasis of dermal structure using in vivo and in vitro wound healing models. Time-course healing of a full-thickness skin punched wound exhibited significantly faster wound closure in Npas2-/- mice than wild-type (WT) C57Bl/6J mice. Dorsal skin fibroblasts isolated from WT, Npas2+/-, and Npas2-/- mice exhibited consistent profiles of core clock gene expression except for Npas2 and Per2. In vitro behavioral characterizations of dermal fibroblasts revealed that Npas2-/- mutation was associated with increased proliferation, migration, and cell contraction measured by floating collagen gel contraction and single-cell force contraction assays. Npas2 knockout fibroblasts carrying sustained the high expression level of type XII and XIV FAICT collagens and synthesized dermis-like thick collagen fibers in vitro. Confocal laser scanning microscopy demonstrated the reconstruction of dermis-like collagen architecture in the wound healing area of Npas2-/- mice. This study indicates that the induced Npas2 expression in fibroblasts may interfere with skin homeostasis, wound healing, and dermal tissue reconstruction, providing a basis for novel therapeutic target and strategy. Anat Rec, 2019. © 2019 Wiley Periodicals, Inc.

Ameliorative effects of curcumin on microvascular complicationin streptozotocin-induced diabetic rat penis

Background: Vascular diseases from diabetes typically result in conditions that interfere with the microvascular functions, such as erectile dysfunction (ED) of the penis. It is unrevealed which morphological changes the microvasculature of the penis may result from the treatment and supplementation of curcumin.Objective: This study aims to determine the effects of curcumin, the active compound of turmeric, on penile microvasculature in experimentally induced diabetic rats.Methods: Rat were divided into three groups, control © and diabetic (DM). Diabetic rats supplemented with curcumin (DMC) (200 mg/kg BW). For induction of a diabetic condition, rats were injected with a single intraperitoneal injection of streptozotocin (60 mg/kg BW). A histological study and vascular corrosion casting along with scanning electron microscopy (SEM) were used to determine a morphological change of microvessels in the penis of rats after 8 weeks of experiments.Results: The histological investigations of penile tissue of DM group revealed thickly lined venous sinuses with smooth but sparse muscle layers. Dorsal arteries (DA) and deep dorsal veins (DV) of the penis were present of narrow lumen and showed thickening of collagen fibers at their wall of tunica media. An increasing thickness of collagen fibers was observed. Vascular corrosion casting revealed critically decreased diameters of DM venous sinus in the penis’ glans, which were observed as shrunken and partially lacking in the distal area of the glans. The diameter of venous sinuses in corpus cavernosum (CC) of DM rats were dilated and the density of venous sinuses was decreased. Interestingly, the signs of vessel restoration and improvement were presented by increasing diameters and recovering with healthy and nourished DA, DV, venous sinus at the area of the glans, and CC of the penis in DMC groups. Conclusion: The results have shown that the effects of curcumin could improve the destruction of the penile microvasculature in induced diabetic rats by STZ. Keywords: Diabetes mellitus, streptozotocin, curcumin, penis, Microvasculature, vascular corrosion cast

The Effect of Using Bone Marrow Mesenchymal Stem Cells Versus Platelet Rich Plasma on the Healing of Induced Oral Ulcer in Albino Rats.

Background and ObjectivesOral ulceration is one of the most common debilitating condition that affects the oral cavity. In this study, the effect of locally injected platelet rich plasma (PRP) and bone marrow-derived mesenchymal stem cells (BMSCs) on the healing of oral ulcer was investigated.Methods and ResultsAn ulcer was induced in buccal mucosa of rats by using 5mm biopsy punch followed by application of cotton swab soaked with formocresol for 60sec. The ulcer was left untreated in the control group, treated with intralesional injection of PRP, or isolated cultured BMSCs. Data were analyzed clinically, histologically and immunohistologically on day 3, 5, 7 and 10. BMSCs group showed smaller ulcer area throughout the whole experimental period than the other groups with complete resolution of the ulcer on day 10, unlike the control group. However, there was no significant difference with PRP, on day 5, 7 and 10, regarding clinical ulcer size. BMSCs group showed better histological results regarding the rate of epithelial cell migration, the number of inflammatory cells, thickness and organization of collagen fibres and the number of blood vessels, with complete re-epithelization on day 10. BMSCs group showed a greater number of anti-PCNA positive nuclei throughout the whole experimental period than the other groups except on day 5, PRP had higher mean numbers of anti-PCNA positive nuclei in both tissues.ConclusionsBoth PRP and BMSCs accelerate wound healing and enhance the quality of the healing tissue with the latter being slightly more effective and faster.

Collagen during odontogenesis and in ameloblastoma: A polarizing microscopic study.

Background:Odontogenesis is a highly coordinated and complex process which depends on cell–cell interactions that result in initiation and generation of tooth. Tissue remnants of developing tooth can form odontogenic tumors possibly, reflecting different developmental stages in tooth formation. In both odontogenesis and odontogenic tumors, stroma plays a prominent role in maintaining epithelial tissues with continuous molecular interactions. As the collagen forms an integral part of connective tissue stroma, in the present study, polarization colors and thickness of the collagen fibers were assessed in both tooth germ papillae and ameloblastoma using picrosirius red (PSR) stain.Materials and Methods:Collagen fibers in 20 cases of ameloblastoma and 10 tooth germs from the human fetus were evaluated with PSR stain and examined under polarizing microscopy.Results:Polarization colors of red-colored collagen fibers with greater diameter were more in ameloblastoma when compared to tooth germ papillae in which green-colored collagen fibers with smaller diameter being more.Conclusion:The absence of hard tissue formation in ameloblastoma might be due to the presence of significantly more number and greater thickness of red-colored collagen fibers. Thus, the nature of collagen fibers can predict the nature in terms of biologic behavior and prognosis.

Role of dental pulp in age estimation: A quantitative and morphometric study.

Context:This study was designed to characterize the role of the dental pulp (DP) in age estimation.Aim:The analysis of age-related quantifiable changes in DP components such as odontoblasts, collagen fibers, and blood vessels.Subjects and Methods:One hundred and twenty extracted teeth from six age groups (20–30 years, 31–40 years, 41–50 years, 51–60 years, 61–70 years, and 71–80 years) were subjected to decalcification and routine histopathological processing followed by Hematoxylin and Eosin and Picrosirius Red staining. Evaluation of the number of odontoblasts, mean vessel area (MVA), mean vessel diameter (MVD), and collagen fiber thickness were done.Statistical Analysis Used:ANOVA test, Fisher's test, and Regression Analysis.Results:Reduction in the number of odontoblasts/mm of pulp-dentinal border, MVA, and MVD were seen with advancing age. Rise in collagen fiber thickness was noted with increasing age. All parameters showed strongly statistically significant differences between age groups with P < 0.001 (ANOVA test).Conclusions:Regression formulae derived for age estimation based on data collected demonstrated linear correlation with age. Collagen fiber thickness had the highest accuracy followed by odontoblast numbers and MVA. MVD was the least accurate among the factors considered. However, the highest accuracy of 90.9% was seen when all parameters were incorporated together in a single equation.