Early unhealthy eating habits underlie morpho-functional changes in the liver and adipose tissue in male rats.

Early-life consumption of high-fat and sugar-rich foods is recognized as a major contributor for the onset of metabolic dysfunction and its related disorders, including diabetes and nonalcoholic fatty liver disease. The lifelong impact of early unhealthy eating habits that start at younger ages remains unclear. Therefore, to better understand the effects of diet, it is essential to evaluate the structural and functional changes induced in metabolic organs and potential mechanisms underlying those changes. To investigate the long-term effects of eating habits, young male rats were exposed to high-sugar and high-energy diets. After 14 weeks, body composition was assessed, and histopathological changes were analyzed in the liver and adipose tissue. Serum biochemical parameters were also determined. Expression of inflammatory markers in the liver was evaluated by immunohistochemistry. Our results revealed that serum levels of glucose, creatinine, aspartate transaminase (AST), alanine transaminase (ALT), and lipid profile were increased in rats red high-sugar and high-energy diets. Histopathological alterations were observed, including abnormal hepatocyte organization and lipid droplet accumulation in the liver, and abnormal structure of adipocytes. In both unhealthy diet groups, hepatic expression of Toll-like receptor 4 (TLR4), cyclooxygenase 2 (COX-2), and E-selectin were increased, as well as a biomarker of oxidative stress. Together, our data demonstrated that unhealthy diets induced functional and structural changes in the metabolic organs, suggesting that proinflammatory and oxidative stress mechanisms trigger the hepatic alterations and metabolic dysfunction.

Conventional histomorphometry and fast free of acrylamide clearing tissue (FACT) visualization of sciatic nerve in chicken (Gallus domesticus).

Histomorphometry and use of the fast free of acrylamide clearing tissue (FACT) protocol were studied on the sciatic nerve in chicken (Gallus domesticus). In the first part of the study, the sciatic nerves of 20 chickens of four age groups (7, 14, 26 and 40 days) were studied (n=5 birds per age class). Their sciatic nerve samples were stained with Hematoxylin and Eosin and Masson's trichrome and were histomorphometrically evaluated. In the second part of the study, FACT protocol was applied on the sciatic nerve of a 26 days old chicken. After clearing of 1.00 mm-thick sciatic nerve sections, they were immunolabelled using Hoechst for nuclei staining and recorded by a Z-stack motorized fluorescent microscope. In the conventional histo-morphometry, the epineurium, perineurium and endoneurium were thicker and the nerve bundle diameter was bigger in the left sciatic nerve of chicken of all age groups compared to the right sciatic nerve. On the contrary, the axon diameter and the myelinated nerve fiber diameter were bigger, the myelin sheath was thicker, the nodes of Ranvier intervals were higher and the density of myelinated nerve fibers was also higher in the right sciatic nerve compared to the left one. In conclusion, histomorphometric parameters in the left and right sciatic nerve during chicken growth were significantly different. Furthermore, the FACT protocol could be used for the 3D imaging of the chicken sciatic nerve and its immunostained evaluation.

Estudio histomorfométrico de folículos tiroideos humanos con microscopía holográfica digital / Histomorphometric study of human thyroid follicles by digital holographic microscopy

Introducción: La microscopía holográfica digital ha permitido a la microscopía óptica hacer uso de herramientas numéricas y computacionales; y esto, a su vez, ha favorecido múltiples avances en el estudio de las células y los tejidos en diferentes campos de la medicina y otras ciencias afines. Objetivo: Describir las características histológicas y morfométricas de los folículos tiroideos humanos con la microscopía holográfica digital. Métodos: Se realizó, desde el punto de vista histomorfométrico, un estudio descriptivo y transversal de folículos tiroideos humanos utilizando una instalación de microscopía holográfica digital. Se empleó la técnica de inclusión en parafina y tinción de hematoxilina-eosina para el procesamiento de las muestras. Se realizaron de 10 a 12 capturas de hologramas por muestra y el método de doble propagación para la reconstrucción de los hologramas. Se calculó el área, el perímetro, el diámetro mayor y menor de los folículos y cavidades foliculares y se realizaron reconstrucciones de imágenes holográficas en tres dimensiones. Se determinó como medida de tendencia central la media aritmética y como medida de dispersión la desviación típica o estándar. Resultados: Parámetros foliculares: área (5140,31 ± 1126,71 µm2); perímetro (2961,54 ± 71,2 µm); diámetro mayor:(921,17 ± 24,34 µm); diámetro menor: (746,67 ± 18,08 µm); altura del epitelio (7,92 ± 0,96). Cavidades foliculares: área (3686,18 ±1023,52 µm2); diámetro mayor: (698,86 ± 19,55 µm) y diámetro menor: (581,15 ± 13,82 µm). Conclusiones: Existen parámetros foliculares, determinados mediante la microscopía holográfica digital, no reportados por la literatura consultada, que resultan de interés en el estudio histológico de los folículos tiroideos humanos(AU)

Introduction: Digital holographic microscopy has made it possible to incorporate the use of numerical and computer tools into optical microscopy. This in turn has led to great progress in the study of cells and tissues in several fields of medicine and related sciences. Objective: Describe the histological and morphometric characteristics of human thyroid follicles using digital holographic microscopy. Methods: A descriptive cross-sectional histomorphometric study was conducted of human thyroid follicles using a digital holographic microscopy facility. Sample processing was based on inclusion technique by paraffin and hematoxylin-eosin staining. Ten to twelve holographic captures were made per sample, and the double propagation method was used for holographic reconstruction. Estimation was carried out of the area, perimeter, and greatest and smallest diameter of follicles and follicular cavities, and tri-dimensional reconstructions were made of holographic images. Arithmetic mean was determined as the measure of central tendency, and typical or standard deviation as the measure of dispersion. Results: Follicular parameters: area (5 140.31 ± 1 126.71 µm2); perimeter (2 961.54 ± 71.2 µm); greatest diameter (921.17 ± 24.34 µm); smallest diameter (746.67 ± 18.08 µm); epithelial height (7.92 ± 0.96). Follicular cavities: area (3 686.18 ± 1 023.52 µm2); greatest diameter (698.86 ± 19.55 µm); smallest diameter (581.15 ± 13.82 µm). Conclusions: A number of follicular parameters determined by digital holographic microscopy have not been reported by the literature consulted, and they are of interest to the histological study of human thyroid follicles(AU)

Estudio histomorfométrico de la corteza cerebelosa con microscopia holográfica digital / Histomorphometric study of the cerebellar cortex with digital holographic microscopy

Se realizó un estudio descriptivo y transversal de la corteza cerebelosa humana, desde el punto de vista histomorfométrico, desde enero hasta septiembre del 2015, con el empleo de la microscopia holográfica digital instalada en el Departamento de Holografía Digital de la Universidad de Oriente en Santiago de Cuba, con vistas efectuar mediciones que permitieran establecer comparaciones con otros estudios. Los cálculos mostraron el grosor de las capas molecular y granulosa, el área, el perímetro, los diámetros mayores y menores del cuerpo y el núcleo de las células de Purkinje. Asimismo, se tomaron imágenes holográficas en tres dimensiones, que posibilitaron concluir la existencia de parámetros determinados mediante este procedimiento, los que no habían sido notificados y que resultan de interés en el estudio histológico de la corteza cerebelosa

A descriptive and cross-sectional study of the human cerebellar cortex, from the histomorphometric point of view, was carried out from January to September, 2015, using the digital holographic microscopy installed in the Digital Holography Department of Oriente University in Santiago de Cuba, aimed at making measurings that allowed to establish comparisons with other studies. The calculations showed the thickness of the molecular and granular layers, the area, perimeter, greatest and smallest diameters of the body and Purkinje cells nucleus. Also, holographic images in three dimensions were taken, that facilitated to conclude the existence of certain parameters by means of this procedure, those that had not been notified and are of interest in the histological study of the cerebellar cortex

Utilidad de la microscopia holográfica digital para el estudio histomorfométrico de eritrocitos humanos / Usefulness of the digital holographic microscopy for the histomorphometric study of human red blood cells

Se realizó una investigación descriptiva y transversal para determinar la utilidad de la microscopia holográfica digital en el estudio histomorfométrico de eritrocitos humanos, mediante el análisis comparativo con la microscopia óptica, en el Departamento de Holografía Digital de la Universidad de Oriente de Santiago de Cuba, de abril del 2014 a igual mes del 2015. A tal efecto se seleccionaron muestras de eritrocitos diluidos en las soluciones amortiguadoras Hepes y Hepa, se emplearon el programa Holodig® y el sistema Matlab® para las mediciones en la técnica holográfica, y en el procesamiento estadístico se aplicó la prueba de la t de Student para la comparación de medias entre ambas técnicas microscópicas. Se obtuvo que la microscopia holográfica digital permitiera observar los límites y contornos de los eritrocitos estudiados y las mínimas variaciones en su forma, además de realizar estudios morfométricos, cuyos resultados mostraron similitud a los obtenidos con la microscopia óptica, y de otros parámetros, como el volumen

A descriptive and cross-sectional investigation to determine the usefulness of the digital holographic microscopy in the histomorphometric study of human red blood cells, by means of the comparative analysis with the optic microscopy, was carried out in the Digital Holography Department of Oriente University in Santiago de Cuba, from April, 2014 to the same month in 2015. To such an effect, red blood cells samples diluted in the Hepes and Hepa shock-absorbing solutions were selected, Holodig program and Matlab® system were used for the measurement in the holographic technique, and in the statistical processing the Student t test was applied for the comparison of means between both microscopic techniques. It was concluded that the digital holographic microscopy allowed to observe the limits and contours of the studied red blood cells and the minimum variations in their form, besides carrying out morphometric studies which results showed similarity to those obtained with the optic microscopy, and of other parameters, as volume

Quantification of Various Inflammatory Cells in Advanced Atherosclerotic Plaques.

INTRODUCTION: Atherosclerosis, the pathological basis of coronary artery disease is being extensively studied as understanding of the complex processes involved in the formation and progression that can provide an insight into prevention and treatment of the same. This is an autopsy study to identify and quantify various inflammatory cells in advanced atherosclerotic plaques. AIM: This study aims at identifying and categorizing the various inflammatory cells present in advanced atherosclerotic plaques, noting their distribution in the plaque, quantifying them using histomorphometry and comparing them across plaques of different AHA types. MATERIALS AND METHODS: Post-mortem angiogram was performed on 3 heart specimens obtained at autopsy of random Road Traffic Accident (RTA) cases which revealed evidence of coronary artery disease. End-arterectomy was done and the arteries with atherosclerotic plaques were cut into serial sections and made into tissue blocks. Sections from these blocks were stained with H & E stain and the plaques were classified based on AHA classification. 50 advanced atherosclerotic plaques of AHA Type IV and V were chosen for this study and were screened for inflammatory cells, first with H & E stain and then with different immunohistochemical stains for T-lymphocytes, B-lymphocytes and neutrophils. The T-lymphocytes thus identified was further sub-typed into CD4+ and CD8+ cells again using IHC markers and the percentage area of each was measured using histomorphometry. Then, these values were compared between AHA Type IV and AHA Type V lesions. RESULTS: It was found that the inflammatory cells found in advanced atherosclerotic plaques were predominantly T-lymphocytes as evidenced by their CD3 positivity and they were found to be distributed mainly around the shoulder region and fibrous cap of the plaque. When categorized further, it was found that CD8+ T-cells were always more than CD4+ T-cells in advanced lesions. Meloperoxidase stain for neutrophils was negative in all the plaques examined. The difference in the amount of inflammatory cells between AHA type IV and Type V was not statistically significant. CONCLUSION: The study of the amount of inflammatory cells in atherosclerotic plaques and understanding their role in the pathophysiology of advanced plaques may have therapeutic implications.

Monetite granules versus particulate autologous bone in bone regeneration.

OBJECTIVE: The aim of this study was to test bone tissue response to monetite granules in comparison with intramembranous autologous bone graft in a rabbit calvaria critical size defect model. MATERIALS AND METHODS: Novel monetite granules were synthesized by thermal conversion of set brushite cement. Eight female New Zealand rabbits were used for this study. Two identical 10mm diameter bicortical cranial defects were created in each animal. One of the defects was grafted with monetite granules while the contralateral was grafted with granules of intramembranous autologous bone as control. Animals were sacrificed 8 weeks after surgery and biopsies were taken for histological and histomorphometrical evaluation under light microscopy. Wilcoxon test was used for statistical analysis. RESULTS: The bone defects treated with either autologous bone or monetite granules were able to heal within the study period. Upon histological observation the defects treated with autologous bone granules resembled the adjacent intact calvaria, whereas the defects treated with monetite showed a high infiltration of new bone and only 13.4±8.4% of remaining granules. The percentage of bone volume in the defects of the control group (71±9%) was 16% higher than in the study group (55±10%) (p<0.05). The percentage of augmented mineralized tissue volume in the study group (68±18%) was not significantly different from the control group (p>0.05). CONCLUSION: The amount of augmented mineralized tissue in the bone defects obtained with monetite granules was not significantly different from that obtained with autologous bone. This study confirms the potential of monetite based biomaterials as an alternative to autologous bone graft.

Histopathologic and histomorphometric studies and determination of IL-8 in patients with periodontal disease.

BACKGROUND: Periodontitis is an inflammatory disease characterized by connective tissue breakdown and alveolar bone resorption. OBJECTIVE: The aim of this study was to make a quantitative evaluation of the gingival tissue components in biopsies from patients with different clinical states of periodontal disease and to determine the relationship between the presence of interleukin-8 and the tissue destruction. MATERIALS AND METHODS: The study group consisted of 33 biopsies from adult subjects. A total of 25 periodontal patients were sub-classified on the basis of the clinical and radiographic criteria in mild, moderate and advanced periodontitis. Gingival samples were obtained from patients in the course of basic periodontal surgeries as a muco-periostic collate. Data were analyzed by the SPSS system. RESULTS: Collagen fibres decreased as non-specific chronic states increased. A lymphocyte inflammatory infiltrate changed to a lymphoplasmocytary form. Reactivity to interleukin-8 was detected with the severity of the histopathologic diagnoses; however there was no association with the clinical diagnoses. CONCLUSIONS: Histopathology and histomorphometry of gingival tissue changed with the degree of inflammation. No evidence of interleukin-8 as a biomarker for clinical diagnoses of periodontitis was obtained.

Porcine study on the efficacy of autogenous tooth bone in the maxillary sinus.

OBJECTIVES: This study sought to elucidate the effect of autogenous tooth bone material by experimenting on minipig's maxillary sinus and performing histological and histomorphometric analyses. MATERIALS AND METHODS: Five 18-24 month-old male minipigs were selected, and right maxillary sinuses were grafted with bone graft material made of their respective autogenous teeth extracted eight weeks earlier. The left sides were grafted with synthetic hydroxyapatite as control groups. All minipigs were sacrificed at 12 weeks after bone graft, which was known to be 1 sigma (σ) period for pigs. Specimens were evaluated histologically under a light microscope after haematoxylin-eosin staining followed by semi-quantitative study via histomorphometric analysis. The ratio of new bone to total area was evaluated using digital software for calculation of area. RESULTS: All specimens were available, except one on the right side (experimental group), which was missing during specimen preparation. This study demonstrated new bone at the periphery of the existing bone in both groups, showing evidence of bone remodeling, however, encroachment of new bone on the central part of the graft at the 1 σ period was observed only in the autogenous tooth bone group (experimental group). Histomorphometric analysis showed more new bone formation in the experimental group compared to the control group. Although the difference was not statistically significant (P>0.05), the mean percentage area for new bone for the experimental and control groups were 57.19%±11.16% and 34.07%±13.09%, respectively. CONCLUSION: The novel bone graft material using autogenous tooth is a good alternative to autogenous bone, comparable to autogenous bone, and outperforming synthetic hydroxyapatite bone graft materials in terms of bone regeneration capacity. Augmentation with autogenous tooth bone materials will reduce donor site morbidity without hampering the safety of the autogenous bone graft.

Alterações da interface de implantes submetidos a carga imediata – estudo experimental em cães Beagle / Implant-bone interface changes due to immediate loading: an experimental study in Beagle dogs

Este estudo in vivo destinou-se a analisar as alterações histológicas que ocorrem na interface osso/implante durante o período crítico após a colocação em carga imediata. O modelo experimental foi desenvolvido na mandíbula de seis cães Beagle, tendo sido colocados dois implantes cilíndricos no terceiro e quarto quadrantes, respetivamente (n=24 implantes). Os implantes no quarto quadrante, em cada animal, foram reabilitados dentro do período de 24 horas, e o outro par, do terceiro quadrante, foi usado como controle, sem reabilitação. As eutanásias ocorreram aos nove, 17 e 31 dias após a colocação em carga, tendo sido preparadas amostras por técnicas não descalcificadas. A relação entre as medições do contato ósseo no terceiro e quarto quadrantes e as forças de mastigação foram analisadas estatisticamente no tempo de experimentação. O contato osso/implante foi ligeiramente maior, sem ser signifi cativo, nos implantes em carga imediata (52,13%), do que nos implantes dos quadrantes utilizados como controle (47,6%). O contato ósseo mostrou uma tendência para um valor menor no segundo grupo, aos 17 dias, e maior no primeiro grupo. Na comparação de contato osso/implante entre os quadrantes, apenas o terceiro grupo apresentou aumento do contato osso/implante no quarto quadrante. Em conclusão, verificamos a presença de atividade osteogênica em todas as interfaces osso/implante e a carga imediata não afetou a osseointegração...

This in vivo study aimed to analyse the histological changes that occur in the bone-implant interface during the critical period following immediate loading. The experimental model was developed in the mandible of six Beagle dogs, having been placed two cylindrical implants in the 3rd and 4th quadrants, respectively (n=24 implants). The 4th quadrant implants, in each animal, were rehabilitated within a period of 24 hours and the other pair, the 3rd quadrant ones, was used as a control without prosthetic rehabilitation. Animals were sacrificed at 9th, 17th and 31st days after loading and samples prepared using non-decalcified techniques. The relationship between bone contact measurement at the 3rd and 4th quadrants, as well as masticatory forces, was statistically analysed in the time of the experiment. The bone-implant contact was slightly higher, without statistical signifi cance, on the immediate loaded implant group (52.13%) than in the quadrants used as controls (47.6%). The bone contact showed a trend towards lower values in the 2nd group (17 days), and higher in the 1st group. When comparing bone-implant contact between quadrants, only the 3rd group showed increased bone-implant contact at the 4th quadrant. In conclusion we found the presence of osteogenic activity in all bone-implant interfaces and immediate loading did not affect osseointegration...

Porcine study on the efficacy of autogenous tooth bone in the maxillary sinus

OBJECTIVES: This study sought to elucidate the effect of autogenous tooth bone material by experimenting on minipig's maxillary sinus and performing histological and histomorphometric analyses. MATERIALS AND METHODS: Five 18-24 month-old male minipigs were selected, and right maxillary sinuses were grafted with bone graft material made of their respective autogenous teeth extracted eight weeks earlier. The left sides were grafted with synthetic hydroxyapatite as control groups. All minipigs were sacrificed at 12 weeks after bone graft, which was known to be 1 sigma (sigma) period for pigs. Specimens were evaluated histologically under a light microscope after haematoxylin-eosin staining followed by semi-quantitative study via histomorphometric analysis. The ratio of new bone to total area was evaluated using digital software for calculation of area. RESULTS: All specimens were available, except one on the right side (experimental group), which was missing during specimen preparation. This study demonstrated new bone at the periphery of the existing bone in both groups, showing evidence of bone remodeling, however, encroachment of new bone on the central part of the graft at the 1 sigma period was observed only in the autogenous tooth bone group (experimental group). Histomorphometric analysis showed more new bone formation in the experimental group compared to the control group. Although the difference was not statistically significant (P>0.05), the mean percentage area for new bone for the experimental and control groups were 57.19%+/-11.16% and 34.07%+/-13.09%, respectively. CONCLUSION: The novel bone graft material using autogenous tooth is a good alternative to autogenous bone, comparable to autogenous bone, and outperforming synthetic hydroxyapatite bone graft materials in terms of bone regeneration capacity. Augmentation with autogenous tooth bone materials will reduce donor site morbidity without hampering the safety of the autogenous bone graft.

Effectiveness of autogenous tooth bone used as a graft material for regeneration of bone in miniature pig

INTRODUCTION: This study examined the effect of autogenous tooth bone used as a graft material for bone regeneration in an artificial bony defect of minipigs. MATERIALS AND METHODS: Four healthy minipigs, weighing approximately 35-40 kg, were used. Four standardized artificial two-walled bony defects, 5 mm in length and depth, were made on the bilateral partial edentulous alveolar ridge on the mandible of minipigs, and autogenous tooth bone was augmented in the right side as the experimental group. On the other hand, only alloplastic bone graft material HA was grafted with the same size and manner in the left side as the control group. All minipigs were sacrificed at 4 weeks after a bone graft and evaluated histologically by Haematoxylin-eosin staining. The specimens were also evaluated semi-quantitatively via a histomorphometric study. The percentage of new bone over the total area was evaluated using digital software for an area calculation. RESULTS: All specimens were available but one in the left side (control group) and two in the right side (experimental group) were missing during specimen preparation. The amount of bone formation and remodeling were higher in all experimental groups than the control. The mean percentage area for new bone in the experimental and control groups was 43.74+/-11.96% and 30.79+/-2.93%, respectively. CONCLUSION: Autogenous tooth bone is a good alternative to autogenous bone with the possible clinical feasibility of an autogenous tooth bone graft in the reconstruction of bony defects.

Histomorphometric Study of Tissue Reaction of Gore-Tex Implant in the Rat Model

To reconstruct the defect caused by losses of bone, cartilage and soft tissue, either autogenous tissue or alloplastic material can be used. Although the autogenous tissue is ideal choice for their reconstruction, the availability and problems associated with donor site morbidity limit its usage. Therefore the alloplastic material have been used widely for tissue replacement. Many synthetic material have been developed for alloplasty; among these, the silicone has been the most popular alloplastic material for plastic surgeon. Inherent problem related to the use of silicone are infection, exposure, implant migration, capsular formation, and poor penetration of bone or soft tissue into the implant. To solve these problems, many effort have been made. From 1970s the porous high density polyethylene(HDPE, Medpor) and expanded polytetrafluorethylene(Gore-Tex), which are porous substance, have been used because those have the advantage of less implant migration by the penetration of connective tissue into the pore, less capsule formation and reduced chance of infection due to high vascularity. The purpose of this study is to compare the tissue reaction of silicone and Medpor with that of Gore-Tex in rats. The 1.0 cm sized square shape plate of silicone, Medpor and Gore-Tex were implanted in the back of rat and bloc tissue specimen were obtained including the implant and their surrounding capsule in 4, 8, 12 weeks after implantation. The specimen were examined grossly and microscopically. The microscopic slide were prepared with H-E and Masson's trichrome stain. Grossly the capsule around the silicone rubber was easily peel off while the capsules around Medpor was strongly adherent to implant and that of Gore-Tex was mildly adherent to implant. Under microscope, the thickness of capsule were compared. 1he capsule of those implants were thicker in 4 weeks then 8, 12 weeks. The capsule of silicone were significant thicker than Medpor and Gore-Tex in 4 weeks (274.8+ 35.1 um vs 119.8+23.2um vs 177.2+27.9 um). Histologically the capsule of silicone and Gore-Tex consist of denser connective tissue and showed lesser foreign body reaction than Medpor. Many multinucleated giant cells were found around the Medpor, while none of them were found around silicone and Gore-Tex. The capsule of Medpor showed more new capillary formation than silicone due to tissue ingrowth into the pore in the implant material so Medpor fixed hardly around the soft tissue and was impossible to remove. But Gore-Tex, that is microporous alloplastic material, have less capsular formation, less foreign body reaction and limited tissue ingrowth and stabilization of implant while at the same time permitting easy removal when necessary. From this study the Gore-tex will be useful alloplastic material in clinical usage by virtue of limited tissue ingrowth and decreased chance of infection, implant migration and capsular formation.