L.P. Nogueira

3D histomorphometric quantification of trabecular bones by computed microtomography using synchrotron radiation.

Conventional bone histomorphometry is an important method for quantitative evaluation of bone microstructure. X-ray computed microtomography is a non-invasive technique, which can be used to evaluate histomorphometric indices in trabecular bones (BV/TV, BS/BV, Tb.N, Tb.Th, Tb.Sp). In this technique, 3D images are used to quantify the whole sample, differently from the conventional one, in which the quantification is performed in 2D slices and extrapolated for 3D case. In this work, histomorphometric quantification using synchrotron 3D X-ray computed microtomography was performed to quantify the bone structure at different skeletal sites as well as to investigate the effects of bone diseases on quantitative understanding of bone architecture. The images were obtained at Synchrotron Radiation for MEdical Physics (SYRMEP) beamline, at ELETTRA synchrotron radiation facility, Italy. Concerning the obtained results for normal and pathological bones from same skeletal sites and individuals, from our results, a certain declining bone volume fraction was achieved. The results obtained could be used in forming the basis for comparison of the bone microarchitecture and can be a valuable tool for predicting bone fragility.

Application of the SR-PhC-μCT technique with phase retrieval for the characterization of internal and external structures of Rhodnius prolixus

The present work is the second one of a two- part publication on the use of Synchrotron Radiation Phase Contrast microtomography (SR-PhC-μCT) for the study of Rhodnius prolixus morphology. In previous paper the visualization of various detailed features were highlighted thanks to the edge enhancement effects typical of the phase contrast technique, but the contrast between foreground and background remained poor. In this study the same data set have been re-visited with application of a single distance phase retrieval algorithm. The resulting slices showed very high quality images that enable a better visualization of important muscles and neurohemal organs of the central nervous system within the head of Rhodnius prolixus.

Mapping lead distribution in bones by dual-energy computed microtomography with synchrotron radiation

We investigated the feasibility of the micro computed tomography dual-energy imaging to map of lead deposition inside bone. Femora of Wistar rats were imaged at the SYRMEP beamline at the ELETTRA synchrotron facility at Trieste, Italy, using two different energies, one just below (15.75 keV) and other just above (16.10 keV) the L-edge absorption energy of lead. By utilizing dual-energy micro-tomography the distribution-map of lead in the specimens was obtained. On the three-dimensional subtraction images, lead distribution could be clearly visualized over trabecular and cortical bone tissues.

Evaluation of rib microstructure in Wistar rats using SR-μCT after radiation therapy simulation for breast cancer

A better understanding of biological interactions that occur after exposure to photon radiation is needed in order to optimize therapeutic regimens and facilitate development and strategies that decrease radiation-induced side effects in humans. In this work, ribs of Wistar rat submitted to radiotherapy simulation were imaged using synchrotron radiation computed microtomography at Elettra Synchrotron Laboratory in Trieste, Italy. Histomorphometric parameters were calculated directly from the 3D microtomographic images and showed significant differences between irradiated and non-irradiated groups.

Analysis of femur head microstructure in ovariectomized rats

It is well accepted that rat ovariectomy (OVX) is a model of estrogen deficiency. OVX played a very important role in the initiating and developing of osteoporosis and it has been shown to be a major risk factor for the development of postmenopausal osteoporosis in women. In this work we used synchrotron radiation computed microtomography to investigate the skeletal effects in femoral head of female Wistar rats after bilateral ovariectomy surgery. The CT system was set up at the SYRMEP beamline in the synchrotron radiation facility ELETTRA (Trieste, Italy). Micro-CT images provided 3D information on precise trabecular microstructure by the reconstruction of multiple 2D images with almost 2 μm resolution. Our aim was to use histomorphometric analysis to reveal the effect of OVX on the three-dimensional (3D) trabecular bone microarchitecture. Evaluated morphometric parameters were trabecular bone volume-tissue volume ratio (BV/TV), trabecular number (Tb.N), trabecular separation (Tb.Sp) and trabecular thickness (Tb.Th). OVX group presented noticeable reduction in the Tb.N and Tb.Th when compared with control group (P 0.05). Our data may help to gain more insight into the potential mechanism of osteoporotic femoral head fractures.

Application of synchrotron radiation computed microtomography for quantification of bone microstructure in human and rat bones

This work aims to evaluate histomorphometric quantification by synchrotron radiation computed microto-mography in bones of human and rat specimens. Bones specimens are classified as normal and pathological (for human samples) and irradiated and non-irradiated samples (for rat ones). Human bones are specimens which were affected by some injury, or not. Rat bones are specimens which were irradiated, simulating radiotherapy procedures, or not. Images were obtained on SYRMEP beamline at the Elettra Synchrotron Laboratory in Trieste, Italy. The system generated 14 μm tomographic images. The quantification of bone structures were performed directly by the 3D rendered images using a home-made software. Resolution yielded was excellent what facilitate quantification of bone microstructures.

Evaluation of photon irradiation treatment upon calcium content of ribs of Wistar rats using micro-XRF

Therapeutic doses of radiation have been shown to have deleterious consequences on bone health. Among the treatment strategies used for breast cancer treatment, the most used are radiotherapy and chemotherapy. Radiotherapy may be given to destroy the cancer cells using high-dose x-rays. Protocols vary considerably, but generally whole body irradiation totals from 10 to 15 Gy, whereas local therapy totals from 40 to 70 Gy. In clinical practice, the quantitative evaluation of bone tissue relies on measurements of bone mineral density values, which are closely associated with the risk of osteoporotic fracture. Improved survivorship rates of cancer patients receiving radiotherapy increase the importance of understanding the mechanisms and long-term effects of radiation-induced bone loss. In this work, we investigated the variation on calcium distribution in ribs of female Wistar rats (Rattus norvegicus) submitted to photon irradiation with a single dose of 20 Gy. The determination of the calcium distribution ...

Synchrotron microtomography to evaluate effects of different polychemotherapy drugs on cortical bone structure

Abstract Purpose: To determine the effects of different polychemotherapy drugs on cortical bone structure, the femur diaphysis of rats were treated with two different chemotherapy drugs, AC (doxorubicin + cyclophosphamide) and TC (docetaxel + cyclophosphamide), and evaluated by 3D morphological analysis using synchrotron radiation microtomography. Materials and methods: Wistar rats were classified into three groups. One group received doses of docetaxel and cyclophosphamide (G1) – TC regimen; a second group received doses of doxorubicin and cyclophosphamide (G2) – AC regimen; while a control group (G0) received no further treatment. 3D tomographic images of the rats’ femurs were obtained at the SYRMEP (Synchrotron Radiation for Medical Physics) beamline at the ELETTRA Synchrotron Laboratory in Trieste, Italy, using monochromatic X-rays with resolution of 9 μm. Results: It could be shown that the treatment caused significant differences in morphological parameters measured from the 3D images of femur diaphysis of rats, among the studied groups, complementing a previous study using stereological methods, biochemistry and electron microscopy. Conclusions: Our results indicate that the same process of osteoporosis caused by advancing age might occur in young women treated with docetaxel + cyclophosphamide (TC) and doxorubicin + cyclophosphamide (AC). 3D microtomography was shown to be an outstanding technique for bone analysis.

Synchrotron radiation X-ray microtomography and histomorphometry for evaluation of chemotherapy effects in trabecular bone structure

Three-dimensional microtomography has the potential to examine complete bones of small laboratory animals with very high resolution in a non-invasive way. One of the side effects caused by some chemotherapy drugs is the induction of amenorrhea, temporary or not, in premenopausal women, with a consequent decrease in estrogen production, which can lead to bone changes. In the present work, the femur heads of rats treated with chemotherapy drugs were evaluated by 3D histomorphometry using synchrotron radiation microcomputed tomography. Control animals were also evaluated for comparison. The 3D tomographic images were obtained at the SYRMEP (SYnchrotron Radiation for MEdical Physics) beamline at the Elettra Synchrotron Laboratory in Trieste, Italy. Results showed significant differences in morphometric parameters measured from the 3D images of femur heads of rats in both analyzed groups.

Analysis of calcium distribution in femur of female rats submitted to different chemotherapy regimens

The most used treatment strategies for breast cancer (BC) are surgery, chemotherapy, and/or radiotherapy. Premenopausal women undergoing adjuvant chemotherapy BC treatment have significant bone loss from the first year after the treatment. This high bone mineral density loss can lead to an increased risk of fractures. Doxorubicin associated with Cyclophosphamide (AC) is a multidrug widely used for BC treatment, although this association can cause severe side effects. Today it is been discussed the use of the Docetaxel and cyclophosphamide (TC) association for BC treatment. The influence on the bone during chemotherapy regimens that include taxanes, like docetaxel, is unknown. Data from the “Women`s Health Initiative” show that postmenopausal BC survivors have a 15% greater risk of developing fractures than women without a history of BC. In this study, it was evaluated parameters involved in osteoporosis when rats were subjected to a chemotherapy regimen (TC) and/or irradiation (IR). Female Wistar rats, 03 months old, were divided into 3 groups: control, TC+IR (G1) and AC+IR (G2). The animals were euthanized 5 after months the end of treatment and their femurs were excised and dissected. Sections of 10 μm thick were used for μXRF analysis at the National Laboratory of Synchrotron Light. The uteri of these rats were collected and weighed Statistical analyzes were performed using GraphPad Prism, and values were compared using ANOVA and Tukey post-hoc test. The obtained results showed that animals from G2 had a significant reduction (p<;0.05) of uterine mass when compared to control. The qualitative analysis performed by μXRF showed that the animals from G2 had iron in bone composition of the femurs. This same result was not observed in animals from G0 and G1 groups. These results suggest that early menopause occurs and osteoporosis begins, probably because of the absence, or reduced, production of estrogen. The presence of iron in the G2 samples indicates the process of osteoporosis, because according to literature, this ion is competitive with calcium ions.