Effect of treatment with simvastatin on bone microarchitecture of the femoral head in an osteoporosis animal model

The objective of this study was to evaluate the microarchitecture and trabecular bone strength at the distal region of the femur, and its biomechanical properties with simvastatin administration with two different doses in ovariectomized (OVX) rats. Ninety rats were divided into six groups to evaluate treatment with the simvastatin drug (n = 15): SH (Sham surgery), SH‐5 (5 mg simvastatin), SH‐20 (20 mg simvastatin), OVX, OVX‐5, and OVX‐20. Euthanasia was performed at three different times, five animals per period: 7, 14, and 28 days. The effectiveness of the treatments was evaluated by mechanical testing and histomorphometric analysis of the femurs. The results of analysis by the linear model of mixed effects showed 20 mg of simvastatin results in increased trabecular bone after 14 days (P = 0.039) of ingestion in ovariectomized animals. However, ingestion of 5 mg of simvastatin is able to sensitize the trabecular bone only at 28 days (P = 0.005) of ingestion. In the mechanical tests stiffness improves within 28 days (P = 0.003). Regarding maximum strength, no statistical differences were observed. According to these results, it can be concluded that for a decrease in oral intake, longer treatment times are required. Microsc. Res. Tech. 79:684–690, 2016. © 2016 Wiley Periodicals, Inc.     

Effects of aerobic training,resistance training,or combined resistance‐aerobic training on the left ventricular myocardium in a rat model

This study follows the left ventricular (LV) hypertrophy in rats undergoing aerobic training alone (A), resistance training alone (R), or combined resistance and aerobic training (RA) (usually referred as concurrent training) program. A sedentary control group (C) was included. LV remodeling was evaluated using electron and light microscopy. The LV weight to body weight (LVW: BW) increased 11.4% in A group, 35% in the R group, and 18% in the RA group compared to the C group. The LV thickness increased 6% in the A group, 17% in the R group, and 10% in the RA group. The LV internal diameter increased 19% in the A group, 3% in the R group, and 8% in the RA group compared with the C group. The cross‐sectional area of cardiomyocyte increased by 1% with the A group, 27% with R group, and 12% with RA training. The capillary density increased by 5.4% with A training, 11.0% with R training, and 7.7% with RA training compared with the C group. The volume fraction of interstitial collagen increased by 0.4% with training A, increased by 2.8% with R training, and 0.9% with RA training. In conclusion, except for the LV internal diameter, which increased more in the A group, the cardiac parameters increased more in the R group than in the other groups and in RA group than in A group. Collagen density increased from 5.4 ± 0.8% in the C group to 5.8 ± 0.6% in the A group (n. s.) (P > 0.05), to 8.2 ± 0.7% in the R group (P < 0.05), and to 6.3 ± 0.4% in the RA group (P < 0.05). These results demonstrate a significant increase for collagen content in the LV with R and RA exercise, but the increase was higher with R training alone than with RA training. Microsc. Res. Tech. 77:727–734, 2014. © 2014 Wiley Periodicals, Inc.     

Ovarian hormones and fasting differentially regulate pituitary receptors for estrogen and gonadotropin‐releasing hormone in rabbit female

To investigate the mechanisms by which caloric restriction affects reproductive function in female rabbits, we measured, in animals intact or ovariectomized (OVX) estrogen‐primed and fed ad libitum or fasted for 48 h, the adenohypophysial expression of estrogen receptor‐alpha (ESR1) and gonadotropin releasing hormone receptor (GnRHR) and the dynamic secretion of LH following GnRH stimulation. Fasting increased the number of GnRHR‐immunoreactive (‐IR) cells in intact animals, whereas reduced the density of ESR1‐IR cells in OVX rabbits. Estrogen priming decreased the number of ESR1‐IR cells in fasted and OVX animals. Ovariectomy increased the number of ESR1‐IR cells in fed rabbits, but caused an opposite effect in both fed and fasted animals treated with estrogen. Fasting down regulated the mRNA levels for ESR1 and GnRHR. Estrogen‐priming reduced the abundance for ESR1 mRNA in both fed and fasted rabbits, and that for GnRHR in fasted rabbits. Ovariectomy halved ESR1 mRNA levels independently of treatment and feeding condition, whereas increased (P < 001) that for GnRHR in estrogen‐primed rabbits. In all rabbits, an LH surge occurred 30 min after GnRH injection but the lowest levels were found in intact fasted rabbits and the highest in fasted, estrogen‐primed animals. The LH profile was similar in intact and OVX rabbits and neither fasting nor estrogen priming modified it. In conclusion, fasting differentially modifies the ESR1 and GnRHR expression in the pituitary, depending on the presence of gonadal hormones, indicating complex interactions between metabolic signals and ovarian steroids. Microsc. Res. Tech. 77:201–210, 2014. © 2013 Wiley Periodicals, Inc.     

Vitamin D₃ attenuates anxiety-like behavior in long-term ovariectomized rats with unpredictable mild stress

The impact of various vitamin D3 (VD3) doses (1.0, 2.5, or 5 mg/kg, s.c.) in mitigating the negative consequences of chronic unpredictable mild stress (CUMS) was investigated. Adult female rats with long-term estrogen deficiency were assessed using the sucrose preference test (SPT), the elevated plus-maze (EPM), the light/dark test (LDT), and the open-field test (OFT) to measure anhedonia-like and anxiety-like behavior. The corticosterone (CS) and adrenocorticotrophic hormone (ACTH) concentrations in blood serum and the brain-derived neurotrophic factor (BDNF) expression in the hippocampus of long-term ovariectomized (OVX) rats were measured by ELISA kits and/ or western blotting. Treatment with VD3 (5.0 mg/kg), similarly to fluoxetine (10.0 mg/kg), significantly reduced the anhedonia profile in the SPT and anxiety-like behavior in the EPM and LDT, and CS and ACTH levels in blood serum. It also elevated BDNF levels in the hippocampus of long-term OVX/CUMS compared to OVX/CUMS/solvent rats. Thus, these findings suggest that VD3 (5.0 mg/kg) administration might attenuate the anxiety-like profile in longterm OVX adult rats subjected to the CUMS. This might occur via activation of the BDNF signaling pathway in the hippocampus and via restoration of CS and ACTH levels in blood serum.     

Effects of chemical agents on physical properties and structure of primary pulp chamber dentin

This study evaluated the effects of chemical agents on the physical properties and structure of primary pulp chamber dentin using surface roughness, microhardness tests, and scanning electron microscopy (SEM). Twenty‐five primary teeth were sectioned exposing the pulp chamber and were divided into five groups (n = 5): NT, no treatment; SH1, 1% sodium hypochlorite (NaOCl); SH1U, 1% NaOCl + Endo‐PTC^®; SH1E, 1% NaOCl + 17% EDTA; and E, 17% EDTA. After dentin treatment, the specimens were submitted to roughness, microhardness testing, and SEM analysis. Roughness and microhardness data were submitted to one‐way ANOVA and Tukey's test (P < 0.05). The SH1E group showed the highest roughness, followed by the E group (P < 0.05) when compared with the NT, SH1, and SH1U groups. Microhardness values of SH1 and SH1U showed no significant difference as compared to the NT (control) group (P > 0.05). Microhardness values could not be obtained in the EDTA groups (SH1E and E). The presence of intertubular dentin with opened dentin tubules was observed in the NT, SH1, and SH1U groups. SH1E showed eroded and disorganized dentin with few opened tubules and the intertubular/peritubular dentin was partially removed. Considering the physical and structural approaches and the chemical agents studied, it can be concluded that NaOCl and NaOCl associated with Endo‐PTC^® were the agents that promoted the smallest changes in surface roughness, microhardness, and structure of the pulp chamber dentin of primary teeth. Microsc. Res. Tech. 77:52–56, 2014. © 2013 Wiley Periodicals, Inc.     

The influence of lead nanoparticles on the morpho‐functional changes of rat liver during the postexposure period

Lead as any heavy metals may be found in soil, water, air, and is used in everyday life. Once in the body, it causes toxic effect, making the liver, which is one of the main organs of detoxification, suffer. Recently, the study of the action of not only ionic forms of lead, but also its nanoparticles, has become topical. The study aims at determining changes in the liver of rats and biochemical changes in their blood both at late term of exposure to nanoparticles of lead compounds and in the post‐exposure period. The study was performed on 120 male rats of Wistar line, which were divided into two series, each series containing four groups. The first and the second groups of animals were intraperitoneally injected with colloidal solution of nanoparticles of lead sulfide of 10 and 30 nm in size, and the third group were intraperitoneally injected with a solution of lead nitrate. The fourth group of animals served as control. In the first series, the investigated substances were administered 60 times within 12 weeks. In the second series, after 60‐fold administration of the investigated substances, the exposure was discontibued and animals were observed for 6 weeks—overall duration of 18 weeks. Histological, morphometrical and biochemical methods were used. The body weight was reduced in the rats exposed to PbS_nano1 at week 12 of experiment and in rats exposed to both PbS_nano1 and Pb(NO₃)₂ in the second series. Absolute liver weight increased at week 12 of experiment in all experimental groups. In the second series this value almost reached that of the control level. Relative liver weight in the animals of all experimental groups was higher than that in the control at week 12 of experiment. In the second series this value remained higher in rats exposed to PbS_nano1. After 12 weeks of exposure dystrophic changes in the liver were found in all experimental groups. At week 6 after the exposure (the second series) destructive changes in the liver decreased. Total protein, albumin, glucose, total lipids, cholesterol, triglycerides content in blood serum corresponded with morphological data. The experiment has demonstrated that the 12 weeks long exposure to lead nanoparticles had harmful effect on the liver. Within the postexposure 6‐weeks period structural changes in the liver and biochemical changes in blood serum decreased. Biochemical changes in blood serum corresponded to the morphological data. By many parameters PbS_nano1 had more pronounced harmful effect. Toxicity of PbS_nano2 and Pb(NO₃)₂ were comparable.     

Comparative study of decalcification versus nondecalcification for histological evaluation of one‐wall periodontal intrabony defects in dogs

Biological reactions between biomaterials and surrounding tissues, analyzed by histology, may be important predictors of clinical healing pattern and selection of slide preparation techniques requires a careful consideration regarding sample properties. In this study, we compared histology of bone specimens prepared with or without decalcification and performed histological and histomorphometrical assessments. For the histological evaluation, one‐wall intrabony defects were created around the mandibular molars of six adult dogs, filled with biphasic calcium phosphate, synthetic bone graft material/recombinant human bone morphogenic protein‐2, and healing pattern was histologically evaluated at 4 and 12 weeks. New bone formation in 5 × 4 × 4 mm defects and the length of new cementum, connective tissue attachments around the teeth and number of osteoclasts were measured by histomorphometric analysis. After decalcification, new cementum was easily observed and was significantly increased at week 4. In nondecalcified samples, significantly increased connective tissue attachments were seen at week 12. After 12 weeks, the number of countable multinucleated osteoclasts was significantly increased by 62% in nondecalcified versus calcified tissue sections (P = 0.030). Histomorphometric results may be significantly affected by histological preparation method and therefore, selecting the most appropriate histological preparation method is essential for reliable diagnosis and evaluation of bony samples in studies analyzing tissue regeneration. Microsc. Res. Tech. 78:94–104, 2015. © 2014 Wiley Periodicals, Inc.     

Comparison of methods for quantifying dental wear caused by erosion and abrasion

Various methods have been applied to evaluate the effect of erosion and abrasion. So, the aim of this study was to check the applicability of stylus profilometry (SP), surface hardness (SH) and focus‐variation 3D microscopy (FVM) to the analysis of human enamel and dentin subjected to erosion/abrasion. The samples were randomly allocated into four groups (n = 10): G1‐enamel/erosion, G2‐enamel/erosion plus abrasion, G3‐dentin/erosion, and G4‐dentin/erosion plus abrasion. The specimens were selected by their surface hardness, and they were subjected to cycles of demineralization (Coca‐Cola®‐60 s) and remineralization (artificial saliva‐60 min). For groups G2 and G4, the remineralization procedures were followed by toothbrushing (150 strokes). The above cycle was repeated 3×/day during 5 days. The samples were assessed using SH, SP, and FVM. For each substrate, the groups were compared using an unpaired t‐test, and Pearson correlation coefficients were calculated (α = 5%). For enamel, both profilometry technique showed greater surface loss when the erosion and abrasion processes were combined (P <0.05). The correlation analysis did not reveal any relationships among SH, SP, and FVM to G2 and G4. There were significant correlation coefficients (–0.70 and –0.67) for the comparisons between the FVM and SH methods in enamel and dentin, respectively, in G1 and G3. Choosing the ideal technique for the analysis of erosion depends on the type of dental substrate. SP was not sufficiently sensitive to measure the effects on dentin of erosion or erosion/abrasion. However, SP, FVM and SH were adequate for the detection of tissue loss and demineralization in enamel. Microsc. Res. Tech., 2013. © 2012 Wiley Periodicals, Inc.     

Effects of sodium hypochlorite as dentin deproteinizing agent and aging media on bond strength of two conventional adhesives

This study evaluated the effect of 10% sodium hypochlorite (NaOCl) as deproteinizing agent and storage media on bond strength (BS) of two etch‐and‐rinse adhesive systems to dentin. Twenty‐eight sound extracted human third molars were divided in four groups (n = 7), according to dentin treatment (conventional etching or etching followed by 10% NaOCl application) and adhesive systems (GB—Gluma 2Bond and OS—One‐Step). After dentin treatments and adhesive application, a composite block was built‐up on dentin surface and teeth were serially sectioned to obtain bonded sticks specimens. The sticks were submitted to three aging conditions: (24H) 24 hr in water (immediate), (SH) 3 hr of NaOCl accelerated‐aging or (1Y) 1 year of water storage. Afterward, submitted to microtensile bond strength test (μTBS), failure modes and adhesive interfaces analyzes. Data were analyzed by two‐way analysis of variance (ANOVA) and Tukey's test (α = .05). Dentin deproteinization before bonding significantly reduced μTBS for GB‐treated group (p < .05), regardless the aging conditions. Water storage for 1 year (1Y) and NaOCl accelerated‐aging (SH) decreased μTBS for both adhesives. Yet, the groups stored in NaOCl (SH) exhibited the lowest BS results (p < .05). Bond strength of deproteinized dentin was dependent on the adhesive system composition and NaOCl accelerated‐aging promoted decreased bond strength and further degradation than water storage for 1 year.     

Effect of medicaments used in endodontic regeneration on the morphological characteristics of bovine radicular dentin: Experimental immature tooth model

The purpose of this study was to evaluate the effect of different combinations of irrigating solutions and intracanal dressings in the pretreatment of bovine radicular dentin, using an experimental immature tooth model. Eighty healthy bovine teeth, simulated with incomplete rhizogenesis, were randomly distributed according to the protocols of root canal dentin pretreatment for a regenerative endodontic procedure (n = 10): Control (irrigation with distilled water); SH (irrigation with 1.5% Sodium Hypochlorite); EDTA (irrigation with 17% EDTA); SH/EDTA (irrigation with 1.5% SH + 17% EDTA); SH/CH/EDTA (irrigation with 1.5% SH + calcium hydroxide paste +17% EDTA); SH/MTAP/EDTA (irrigation with 1.5% SH + modified triple antibiotic paste + EDTA 17%); SH/TAP/EDTA (irrigation with 1.5% SH + triple antibiotic paste +17% EDTA) and SH/DAP/EDTA (irrigation with 1.5% SH + double antibiotic paste + EDTA 17%). After the completion of the protocol, the demineralization, the exposure of collagen fibers, and the dentin erosion was evaluated under scanning electron microscopy (SEM), by applying a score system (1–3) to classify the observed features. Statistical analysis was performed (Kruskal‐Wallis and Dunn Multiple Comparison tests—p < .05). SH/TAP/EDTA and SH/DAP/EDTA groups presented the highest rates of demineralization in both the coronal and middle thirds of the root (p < .05). In the SH/MTAP/EDTA group, the samples presented moderate demineralization. The samples from the SH/CH/EDTA group presented similar findings to the control group (p < .05). Conventional triple antibiotic (TAP) and double antibiotic (DAP) pastes promoted more pronounced morphological changes on the dentin surface.     

Evaluation of testicular tissue of adult rats treated with cisplatin incorporated into the liposome

Cisplatin (CPL) is one of the most widely used and effective chemotherapeutic agents for the treatment of several human malignancies. However, it causes serious side effects, especially on reproduction. In order to reduce the undesirable effects caused by many drugs, liposomes have been used as a good system for drug delivery. The aim of this study was to investigate, for the first time, the effects of CPL incorporated into the dipalmitoyl phosphatidylcholine liposome (DPPC) on the testicular tissue of adult Wistar rats. The animals (n = 20) were distributed into four experimental groups: (a) control (distillated water); (b) liposome (DPPC, 1 mL), (c) cisplatin incorporated into liposome (CPL/DPPC), and (d) CPL (8 mg/kg body weight). The animals received a single intraperitoneal injection and were killed 10 days after each treatment for histopathological analysis of testes. The results showed that the testicular histomorphometric parameters in rats of DPPC and CPL/DPPC groups were similar to those of the control group. Meanwhile, rats of the CPL‐treated group showed a variety of morphological alterations, including atrophy of seminiferous tubules and presence of multinucleated cells in the germinal epithelium. The incorporation of CPL into the liposome had no influence on the testicular weight or any other stereological parameters, but it was beneficial in maintaining the body weight of the animals. In conclusion, the liposome suppressed the cytotoxic effects caused by cisplatin in the testes of rats, suggesting a possible use in chemotherapy against cancer to reduce the side effects seen on reproduction. Microsc. Res. Tech. 78:323–329, 2015. © 2015 Wiley Periodicals, Inc.     

Spatial and temporal changes of subchondral bone proceed to articular cartilage degeneration in rats subjected to knee immobilization

This study was aimed to investigate the spatial and temporal changes of subchondral bone and its overlying articular cartilage in rats following knee immobilization. A total of 36 male Wistar rats (11–13 months old) were assigned randomly and evenly into 3 groups. For each group, knee joints in 6 rats were immobilized unilaterally for 1, 4, or 8 weeks, respectively, while the remaining rats were allowed free activity and served as external control groups. For each animal, femurs at both sides were dissected after sacrificed. The distal part of femur was examined by micro‐CT. Subsequently, femoral condyles were collected for further histological observation and analysis. For articular cartilage, significant changes were observed only at 4 and 8 weeks of immobilization. The thickness of articular cartilage and chondrocytes numbers decreased with time. However, significant changes in subchondral bone were defined by micro‐CT following immobilization in a time‐dependent manner. Immobilization led to a thinner and more porous subchondral bone plate, as well as a reduction in trabecular thickness and separation with a more rod‐like architecture. Changes in subchondral bone occurred earlier than in articular cartilage. More importantly, immobilization‐induced changes in subchondral bone may contribute, at least partially, to changes in its overlying articular cartilage. Microsc. Res. Tech. 79:209–218, 2016. © 2016 Wiley Periodicals, Inc.     

Comparative effects of two different doses of low‐level laser therapy on wound healing third‐degree burns in rats

Burns are injuries caused by direct or indirect contact to chemical, physical, or biological agents. Low‐level laser therapy (LLLT) is a promising treatment since it is low‐cost, non‐invasive, and induces cell proliferation. This study aimed to investigate the effects of LLLT (660 nm) at two different fluences (12.5 J/cm² and 25 J/cm²) per point of application on third‐degree burns in rats. Thirty rats (Wistar) divided into GC, GL12.5, and GL25 were used in the study, and submitted to burn injury through a soldering iron at 150°C, pressed on their back for 10 s. LLLT was applied immediately, and 2, 4, 6, and 8 days after wound induction. Histological analysis revealed a decreased inflammatory infiltrate in the group treated with 25 J/cm², and intense inflammatory infiltrate in the control group and in the group treated with 12.5 J/cm². The immunostaining of COX‐2 was more intense in the control groups and in the group treated with 12.5 J/cm² than in the group treated with 25 J/cm². Conversely, VEGF immunomarking was more expressive in the group treated with 25 J/cm² than it was in the other two groups. Therefore, our findings suggest that the use of 25 J/cm² and 1 J of energy was more effective in stimulating the cellular processes involved in tissue repair on third‐degree burns in rats by reducing the inflammatory phase, and stimulating angiogenesis, thus restoring the local microcirculation which is essential for cell migration. Microsc. Res. Tech. 79:313–320, 2016. © 2016 Wiley Periodicals, Inc.     

The effects of human menstrual blood stem cells‐derived granulosa cells on ovarian follicle formation in a rat model of premature ovarian failure

Many studies have reported that human endometrial mesenchymal stem cells (HuMenSCs) are capable of repairing damaged tissues. The aim of the present study was to investigate the effects of HuMenSCs transplantation as a treatment modality in premature ovarian failure (POF) associated with chemotherapy‐induced ovarian damage. HuMenSCs were isolated from menstrual blood samples of five women. After the in vitro culture of HuMenSCs, purity of the cells was assessed by cytometry using CD44, CD90, CD34, and CD45 FITC conjugate antibody. Twenty‐four female Wistar rats were randomly divided into four groups: negative control, positive control, sham, and treatment groups. The rat models of POF used in our study were established by injecting busulfan intraperitoneally into the rats during the first estrus cycle. HuMenSCs were transplanted by injection via the tail vein into the POF‐induced rats. Four weeks after POF induction, ovaries were collected and the levels of Amh, Fst, and Fshr expression in the granulosa cell (GC) layer, as well as plasma estradiol (E2) and progesterone (P4) levels were evaluated. Moreover, migration and localization of DiI‐labeled HuMenSCs were detected, and the labeled cells were found to be localized in GCs layer of immature follicles. In addition to DiI‐labelled HuMenSCs tracking, increased levels of expression of Amh and Fshr and Fst, and the high plasma levels of E2 and P4 confirmed that HuMenSC transplantation had a significant effect on follicle formation and ovulation in the treatment group compared with the negative control (POF) group.     

Decrease of intestinal tumors induced by 1,2-dimethylhydrazine in rats fed with cow milk and buffalo milk

Epidemiological studies in human beings and experimental studies in laboratory animals suggest that milk and dairy products can inhibit effects on the development of some kinds of tumors. Cow milk contains sphingomyelin, butyric acid, conjugated linoleic acid, calcium, vitamin A, carotene and vitamin D. All of these components are known to inhibit the process of carcinogenesis. Our objective was to determine the effect of cow milk and water buffalo milk on the development of colon neoplasias in an experimental model of carcinogenesis in rats induced with 1,2-dimethylhydrazine (DMH). Three-month-old Wistar male rats with an average body weight of 180 g were given a nutritionally adequate diet and drinking water adlivitum, cow milk or water buffalo milk. The milk diets were provided two weeks before the first DMH treatment and their administration was continued during the 10 weeks of DMH treatment. Milk administration finished two weeks after the last DMH doses treatment. Four months after the last carcinogen injection, all surviving animals were sacrificed and examined for intestinal tumors. The number, size, and location of the tumors were recorded and gross pathology was described. Small tumors (< 2.5 mm) were examined by Scanning Electron Microscopy (SEM). Significantly fewer tumors were observed in both groups treated with DMH and supplemented with milk, than in the group treated with DMH without milk administration.     

Creatine supplementation in trained rats causes changes in myenteric neurons and intestinal wall morphometry

Creatine is widely used by athletes as an ergogenic resource. The aim of this study was to evaluate the influence of creatine supplementation on the duodenum of rats submitted to physical training. The number and myenteric neuronal cell bodies as well mucosal and muscular tunic morphometry were evaluated. Control animals received a standard chow for 8 weeks, and the treated ones received the standard chow for 4 weeks and were later fed with the same chow but added with 2% creatine. Animals were divided in groups: sedentary, sedentary supplemented with creatine, trained and trained supplemented with creatine. The training consisted in treadmill running for 8 weeks. Duodenal samples were either processed for whole mount preparations or for paraffin embedding and hematoxylin-eosin staining for histological and morphometric studies of the mucosa, the muscular tunic and myenteric neurons. It was observed that neither creatine nor physical training alone promoted alterations in muscular tunic thickness, villus height or crypts depth, however, a reduction in these parameters was observed when both were associated. The number of myenteric neurons was unchanged, but the neuronal cell body area was reduced in trained animals but not when training and creatine was associated, suggesting a neuroprotector role of this substance .     

Study of the osteoconductive capacity of hydroxyapatite implanted into the femur of ovariectomized rats

Osteoporosis is a global public health that affects postmenopausal women due to the deficiency of estrogen, a hormone that plays an important role in the microarchitecture of bone tissue. Osteoporosis predisposes to pathological bone fracture that can be repaired by conventional methods. However, depending on the severity and quantity of bone loss, the use of autogenous grafts or biomaterials such as hydroxyapatite might be necessary. The latter has received increasing attention in the medical field because of its good biological properties such as osteoconductivity and biocompatibility with bone tissue. The objective of this study was to evaluate using histologic and radiographic analyses, the osteogenic capacity of hydroxyapatite implanted into the femur of rats with ovariectomy-induced osteoporosis. Eighteen rats were divided into three groups with six animals in each: group nonovariectomized, bilaterally ovariectomized not receiving estrogen replacement therapy, and bilaterally ovariectomized submitted to estrogen replacement therapy. Defects were created experimentally in the distal epiphysis of the femur with a surgical drill and filled with porous hydroxyapatite granules. The animals were sacrificed 8 weeks after surgery. The volume of newly formed bone in the implant area was quantified by morphometrical methods. The results were analyzed by ANOVA followed by the Tukey test (P < 0.05). The hydroxyapatite granules showed good radiopacity. Histological analysis revealed less quantity of newly formed bone in the ovariectomized group not submitted to hormone replacement therapy. In conclusion, bone neoformation can be expected even in bones compromised by estrogen deficiency, but the quantity and velocity of bone formation are lower.     

Measuring microenvironment mechanical stress of rat liver during diethylnitrosamine induced hepatocarcinogenesis by atomic force microscope

We developed a highly sensitive method to detect liver tissue stiffness with atomic force microscopy (AFM), and investigated the physical features of hepatocarcinogenesis. Wistar rats received weekly intraperitoneal injections of diethylnitrosamine (DEN) or saline (control) followed by a 2-week wash-out period. Liver samples were harvested at 10, 14, or 18 weeks for pathological examination and stress detection. Previously normal liver tissues developed fibrosis and carcinoma after DEN administration. Although the elastic modulus (E) values of the normal (saline; 0.18 ± 0.04 MPa), fibrotic (8 weeks DEN; 0.25 ± 0.06 MPa) and cirrhotic (12 weeks DEN; 0.39 ± 0.06 MPa) tissues were significantly different, there was no significant difference between the E values of the cirrhotic and the hepatic cell carcinoma (16 weeks DEN; 0.42 ± 0.07 MPa) tissues. Thus, tissue stiffness quantitatively increases during hepatocarcinogenesis, and AFM can be used to sensitively and precisely detect liver stiffness at the microscopic level. Microsc. Res. Tech. 2009. © 2009 Wiley-Liss, Inc.     

Bone repair investigation using rhBMP-2 and angiogenic protein extracted from latex

Background: The aim of this work was to study the new bone tissue formation after bone morphogenetic protein type 2 (rhBMP‐2) and P‐1 application, using 5 and 10 μg of each, combined to a material carrier, in critical bone defects. Methods: It was used 70 Wistar rats (male, ～250 g) that were divided in 10 groups with seven animals on each. Groups are the following: critical bone defect only, pure monoolein gel, 5 μg of pure P‐1, 5 μg of pure rhBMP‐2, 5 μg of P‐1/monoolein gel, 5 μg of rhBMP‐2/monoolein gel, 10 μg of pure P‐1, 10 μg of pure rhBMP‐2, 10 μg of P‐1/monoolein gel, 10 μg of rhBMP‐2/monoolein gel. Animals were sacrificed after 4 weeks of the surgical procedure and the bone samples were submitted to histological, histomorphometrical, and immunohistochemical evaluations. Results: Animals treated with pure P‐1 protein, in both situations with 5 μg and 10 μg, had no significant difference (P > 0.05) for new bone formation; other groups treated with 10 μg were statistically significant (P < 0.05) among themselves and when compared with groups in which it was inserted the monoolein gel or critical bone defect only (P < 0.05). In the group involving the 10 μg rhBMP‐2/monoolein gel association, it was observed an extensive bone formation, even when compared with the same treatment without the gel carrier. Conclusion: Using this experimental animal model, more new bone tissue was found when it was inserted the rhBMP‐2, especially when this protein was combined to the vehicle, and this process seems to be dose dependent. Microsc. Res. Tech., 2011.© 2011 Wiley Periodicals, Inc.