Immunohistochemical and Morphometric Assessment on the Biological Function and Vascular Endothelial Cells in the Initial Process of Cortical Porosity in Mice With PTH Administration.

To clarify the cellular mechanism of cortical porosity induced by intermittent parathyroid hormone (PTH) administration, we examined the femoral cortical bone of mice that received 40 µg/kg/day (four times a day) human PTH (hPTH) (1-34). The PTH-driven cortical porosity initiated from the metaphyseal region and chronologically expanded toward the diaphysis. Alkaline phosphatase (ALP)-positive osteoblasts in the control mice covered the cortical surface, and endomucin-positive blood vessels were distant from these osteoblasts. In PTH-administered mice, endomucin-reactive blood vessels with TRAP-positive penetrated the ALP-positive osteoblast layer, invading the cortical bone. Statistically, the distance between endomucin-positive blood vessels and the cortical bone surface abated after PTH administration. Transmission electron microscopic observation demonstrated that vascular endothelial cells often pass through the flattened osteoblast layer and accompanied osteoclasts in the deep region of the cortical bone. The cell layers covering mature osteoblasts thickened with PTH administration and exhibited ALP, α-smooth muscle actin (αSMA), vascular cell adhesion molecule-1 (VCAM1), and receptor activator of NF-κB ligand (RANKL). Within these cell layers, osteoclasts were found near endomucin-reactive blood vessels. In PTH-administered femora, osteocytes secreted Dkk1, a Wnt inhibitor that affects angiogenesis, and blood vessels exhibited plasmalemma vesicle-associated protein, an angiogenic molecule. In summary, endomucin-positive blood vessels, when accompanied by osteoclasts in the ALP/αSMA/VCAM1/RANKL-reactive osteoblastic cell layers, invade the cortical bone, potentially due to the action of osteocyte-derived molecules such as DKK1.

Idiopathic Avascular Necrosis of the Capitate Treated with a Medial Femoral Trochlea Vascularized Osteochondral Flap: A Case Report.

CASE: Capitate avascular necrosis should be entertained in a differential diagnosis of young, active adults with midcarpal wrist pain. We present a case study of a 30-year-old laborer who developed avascular necrosis (AVN) of his right proximal capitate. Grip strength and wrist motion were limited on examination, with advanced imaging confirming AVN. A diagnostic arthroscopy confirmed the pathology. Treatment was completed with a medial femoral trochlea vascularized flap for cartilaginous resurfacing. At 10-month follow-up, the patient's capitate was healed with stable fixation, and he is working full-time as a laborer without restrictions. CONCLUSION: AVN of the capitate is a unique and challenging articular pathology that requires a thoughtful preoperative evaluation and meticulous surgical technique to reconstruct. The medial femoral trochlea (MFT) vascularized bone transfer with cartilaginous resurfacing is 1 available treatment option. This flap is harvested from the medial femur using microsurgical techniques, based on the descending genicular artery. Using a 2-surgeon approach, simultaneous dissection of the AVN is completed at the wrist. This flap is a vascularized option that can be used for both AVN and nonunion with structural deformity before salvage surgeries.

Fracture risk assessment of vascularized medial femoral condylar bone graft: A finite element analysis.

BACKGROUND: Vascularized medial femoral condyle (MFC) bone graft is useful for pseudarthrosis and osteonecrosis, but has the risk of fracture as a complication. This study aimed to create multiple three-dimensional (3D) finite element (FE) femur models to biomechanically evaluate the fracture risk in the donor site of a vascularized MFC bone graft. METHODS: Computer tomography scans of the femurs of nine patients (four males and five females) with no left femur disease were enrolled in the study. A 3D FE model of the left femur was generated based on the CT images taken from the patients. The descending genicular artery (DGA), the main nutrient vessel in vascularized MFC bone grafts, divides into the proximal transversal branch (TB) and the distal longitudinal branch (LB) before entering the periosteum. Thirty-six different bone defect models with different sizes and locations of the harvested bone were created. RESULTS: The highest stress was observed in the proximal medial and metaphyseal portions under axial and external rotation, respectively. In the bone defect model, the stress was most elevated in the extracted region's anterior or posterior superior part. Stress increased depending on proximal location and harvested bone size. CONCLUSION: Increasing the size of the bone graft proximally raises the stress at the site of bone extraction. For bone grafting to non-load-bearing areas, bone grafting distally using LB can reduce fracture risk. If TB necessitates a larger proximal bone extraction, it is advisable to avoid postoperative rotational loads.

Synovium and blood capillaries at the femoral attachment of anterior cruciate ligament are significantly abundant than those at the tibial attachment / Sinovial y capilares sanguíneos en la inserción femoral del ligamento cruzado anterior son significativamente más abundantes que los de la inserción tibial

SUMMARY: The anterior cruciate ligament (ACL) is a ligament that mainly controls the anterior and rotational mobility of the knee joint, and its surface is covered by a synovial membrane with large number of blood vessels. In general, nutritional supply to the ligament is from many capillaries in the adjacent synovium. However, statistical studies of the capillaries distributed to the ACL are insufficient. In this study, we examined cross-sectional histological images of the femoral attachment (femoral level), middle level of the tendon (middle level), and tibial attachment (tibial level) of the ACL and statistically analyzed blood capillary distribution among the three levels. The ACLs of 10 cadavers were divided into 5 equal sections, and 4mm-thick paraffin sections were made at the femoral level, middle level, and tibial level, and then hematoxylin-eosin (HE) staining were performed. The area of each transverse section was measured using Image-J 1.51n (U. S. National Institutes of Health, Bethesda, MD, USA). Fiber bundles of the ACL were relatively small and sparse in cross-sectional area at the femoral level and became larger and denser toward the tibial level. Many blood levels. The synovium at the attachment of ACL covered the surface of the fiber bundle and also penetrated deeply between the fiber bundles. In particular, the blood capillaries were densely distributed in the synovium at the femoral attachment rather than another two levels. Indeed, the number of capillaries were also most abundant in the femoral level. The cross-sectional ACL area at the femoral level is significantly small, however, the blood capillaries were most abundant. Therefore, when the ACL is injured, its reconstruction with preservation of the femoral ligamentous remnant may be clinically useful for remodeling of the grafted tendon.

El ligamento cruzado anterior (LCA) es un ligamento que controla principalmente la movilidad anterior y rotacional de la articulación de la rodilla, y su superficie está cubierta por una membrana sinovial con gran cantidad de vasos sanguíneos. En general, el suministro de nutrientes al ligamento proviene de muchos capilares en la sinovial adyacente. Sin embargo, los estudios estadísticos de los capilares distribuidos en el LCA son insuficientes. En este estudio, examinamos imágenes histológicas trans- versales de la inserción femoral (nivel femoral), el nivel medio del tendón (nivel medio) y la inserción tibial (nivel tibial) del LCA y analizamos estadísticamente la distribución de los capilares sanguíneos entre los tres niveles. Los LCA de 10 cadáveres se dividieron en 5 secciones iguales y se realizaron cortes en parafina de 4 µm de espesor a nivel femoral, medio y tibial, y luego se realizó tinción con hematoxilina-eosina (HE). El área de cada sección transversal se midió utilizando Image-J 1.51n (Institutos Nacionales de Salud de EE. UU., Bethesda, MD, EE. UU.). Los haces de fibras del LCA eran relativamente pequeños y escasos en el área de la sección transversal a nivel femoral y se hicieron más grandes y más densos hacia el nivel tibial. La membrana sinovial en la unión del LCA cubría la superficie del haz de fibras y también penetraba profundamente entre entre los haces de fibras. En particular, los capilares sanguíneos estaban densamente distribuidos en la unión femoral de la sinovial respecto a los otros dos niveles. De hecho, el número de capilares también fue más abundante a nivel femoral. El área transversal del LCA a nivel femoral era significativamente pequeña, sin embargo, los capilares sanguíneos fueron los más abundantes. Por lo tanto, cuando hay una lesión del LCA su reconstrucción con preservación del ligamento femoral remanente puede ser clínicamente útil para remodelar el tendón injertado.

Copper-containing titanium alloys promote angiogenesis in irradiated bone through releasing copper ions and regulating immune microenvironment.

Poor vascularization was demonstrated as a factor inhibiting bone regeneration in patients receiving radiotherapy. Various copper-containing materials have been reported to increase angiogenesis, therefore might improve bone formation. In this study, a Ti6Al4V-1.5Cu alloy was prepared using selective laser melting (SLM) technology. The immunomodulatory and pro-angiogenic effects of the Ti6Al4V-1.5Cu alloys were examined. In vitro, Ti6Al4V-1.5Cu stimulated vascular formation by restraining inflammatory factors and provoking angiogenic factors in non-irradiated and irradiated macrophages. In vivo, the angiogenic effects of the Ti6Al4V-1.5Cu alloy were confirmed using an irradiated rat femur defect model. Moreover, we found that the biological effects of the Ti6Al4V-1.5Cu alloy were partially due to the release of copper ions and associated with PI3K-Akt signaling pathway. In conclusion, this study indicated the potential of the Ti6Al4V-1.5Cu alloy to promote angiogenesis by releasing copper ions and inhibiting inflammation in normal and irradiated tissues.

Anatomical analysis of different helical plate designs for distal femoral fracture fixation.

PURPOSE: Helical plates potentially avoid the medial neurovascular structures of the thigh. Two implant designs for additional medial 90° helical plate in double plate constructs for geriatric patients and 180° helical plate for single plating in young patients are potential alternatives to widely used standard straight plates. AIMS: (1) assess the distances to adjacent anatomical structures being at risk when applying medial 90° and 180° helical plates with MIPO technique to the femur, (2) compare these distances with medial straight plates, and (3) correlate measurements performed during surgical dissection with CT angiography. METHODS: MIPO was performed in ten human cadaveric femoral pairs using either a 90° helical 14-hole LCP (Group 1) or a 180° helical 15-hole LCP-DF (Group 2). Using CT angiography, distances between femoral arteries and plates as well as distances between plates and perforating vessels were evaluated. Following, specimens were dissected and distances determined again. All plates were removed and measurements were repeated with straight medial plates (Group 3). RESULTS: Overall closest distances between plates and femoral arteries were 14.5 mm (11-19 mm) in Group 1, 21.6 mm (15-24 mm) in Group 2 and 6.5 mm (5-8 mm) in Group 3, with significant differences between Group 3 and both other groups (p < 0.001). Distances to the nearest perforating vessels were 22.4 mm (15-30 mm) in Group 1 and 1.2 mm (1-2 mm) in Group 2. Measurement techniques (visual after surgical disection and CT angiography) demonstrated a strong correlation (p < 0.010). CONCLUSIONS: Inserting 90° and 180° helical plates with MIPO technique is safe, however, attention must be paid to the medial neurovascular structures with 90° helical plates and to the proximal perforating vessels with 180° helical plates. Helical plates can avoid irritation of medial neurovascular structures - compared with straight plates - although care must be taken during their distal insertion. Measurements during surgical dissection correlate with CT angiography.

Mesenchymal stem cells prevent ovariectomy-induced osteoporosis formation in mice through intraosseous vascular remodeling.

Mesenchymal stem cells (MSCs) can promote osteogenesis and are a promising therapy for postmenopausal osteoporosis. However, the relationship between improved intraosseous microcirculation and increased bone mass induced by MSCs in postmenopausal osteoporosis remains unclear. After the primary MSCs were characterized, they were transplanted into ovariectomized mice. MSCs transplantation enhanced the trabecular number, trabecular bone volume/total volume, and trabecular bone mineral density in ovariectomized mice. To determine the role of MSCs in vascular repair, mice were subjected to femoral artery ligation. Through laser speckle flowmetry, vascular perfusion and femoral trabecular bone and cortical bone analyses, we determined the effects of MSCs in promoting intraosseous angiogenesis and preventing osteoporosis in mice. MSCs effectively prevented postmenopausal osteoporosis development, which is associated with the involvement of MSCs in reestablishment of microcirculation within the skeleton.

Quantitative analysis of distal femoral epiphysis blood supply in healthy infants with superb microvascular imaging: a pilot study

Background/aim: This study aimed to determine the blood supply of the distal femoral epiphysis (DFE) using superb microvascular imaging in newborns and infants, and to investigate the correlation with ossification center (OC) length, sex, and age. Materials and methods: A total of 140 cases were evaluated in this study. The cases were divided into 2 groups of less than 90 days and over 90 days. Cartilage blood supply was measured with vascularity index (%) (VI). Results: The mean OC length and median VI values were measured as 10.20 ± 3.72 mm and 0.80% (0.58­1.50) for boys and 10.03 ± 3.36 mm and 0.70% (0.30­1.40) for girls, respectively. There was no significant difference in OC length and VI between sexes. The mean OC length in Group II was significantly higher than in Group I (12.14 ± 3.14 vs 8.09 ± 2.64) (p < 0.001). The median VI in Group I was higher than in Group II (1.40% vs 0.40%) (p < 0.001). There were positive correlations between age and OC length (r = 0 .716), negative correlations between age and VI (r = ­0.822), and between VI and OC length (r = ­0.657). Conclusion: Quantitative reference values for DFE blood supply and OC length can guide the diagnosis and follow-up of many skeletal diseases.

Medial femoral trochlea flap reconstruction: Clinical outcomes and perspectives.

INTRODUCTION: We report our experience with use of the medial femoral trochlea (MFT) osteochondral flap for carpal applications. METHODS: Outcomes of all patients treated with MFT flaps were reviewed. Healing, range of motion, grip strength, carpal alignment, pain, and complication data were collected. RESULTS: MFT flaps were performed on seven patients with a mean age of 26.1 (range, 17-42) years. Indications included scaphoid proximal pole nonunion (n = 3), Kienböck's disease (n = 3), and Preiser's disease (n = 1). The mean follow-up was 32.3 (range, 5-70) months. Union was achieved in five patients at a mean of 12 (range 6-22) weeks. All five patients had increased grip strength and absence of pain at follow-up. There were two failures due to graft resorption. CONCLUSIONS: The MFT osteochondral flap is a technically challenging yet powerful tool to replace the loss of both carpal articular cartilage and adjacent bone.

Low-activity programming of the PDGFRß/FAK pathway mediates H-type vessel dysplasia and high susceptibility to osteoporosis in female offspring rats after prenatal dexamethasone exposure.

Dexamethasone is a common synthetic glucocorticoid drug that can promote foetal lung maturity. An increasing number of studies have shown that prenatal dexamethasone exposure (PDE) can cause a variety of short-term and long-term hazards to offspring, including bone development toxicity. H-type vessels are a newly discovered subtype of blood vessels associated with promoted bone formation and maintenance of bone mass. In this study, we aimed to explore whether H-type blood vessels are involved in PDE-induced long bone development toxicity in offspring and its mechanism. In vivo, we injected dexamethasone (0.2 mg/kg.d) subcutaneously at gestational days 9-20 and observed the H-type vessel abundance and bone mass at different time points in the offspring rats. In vitro, we investigated the effect of dexamethasone (0, 20, 100, and 500 nM) on the tube formation function of rat bone marrow-derived endothelial progenitor cells (EPCs) and explored its mechanism. Our results showed that the adult PDE female offspring rats were susceptible to osteoporosis. In addition, PDE inhibited bone mass, H-type vessel formation and the expression of bone platelet-derived growth factor receptor ß (PDGFRß)/focal adhesion kinase (FAK) pathway-related genes in antenatal and postnatal female offspring. Moreover, PDE promoted the expression of bone glucocorticoid receptor (GR), CCAAT and enhancer binding protein α (C/EBPα) and miR-34c in female foetuses. Dexamethasone suppressed the tube formation of rat bone marrow-derived EPCs and the activity of the PDGFRß/FAK pathway, which was mediated by GR/C/EBPα/miR-34c signalling activation. In summary, PDE can cause H-type vessel dysplasia and high susceptibility to osteoporosis in female offspring, and its mechanism is related to the low-activity programming of the PDGFRß/FAK pathway induced by GR/C/EBPα/miR-34c signalling activation. This study enhances the understanding of the molecular mechanism of dexamethasone-induced bone development toxicity and provides new insights for exploring the early intervention and therapeutic targets of foetal-derived osteoporosis.

Anatomic Relationship of the Femoral Neurovascular Bundle in Patients With Congenital Femoral Deficiency.

BACKGROUND: Congenital femoral deficiency (CFD) is a rare condition that affects the morphology of the hip and surrounding soft tissues. Bony deformity and distorted muscular anatomy are well known, but no studies have described the relationship of the femoral neurovascular (NV) bundle to surgically relevant anatomic landmarks. The authors compared the location of the femoral NV bundle on the affected side in patients with CFD with the unaffected side. The authors hypothesized that the bundle on the pathologic side would be in an abnormal position relative to the unaffected side. METHODS: Thirty-three patients diagnosed with unilateral CFD who had undergone preoperative magnetic resonance imaging of the pelvis were included in our study. The authors identified the femoral NV bundle on the axial cuts and measured its distance from the anterior superior iliac spine (ASIS), anterior inferior iliac spine (AIIS), and lesser trochanter (LT). Anatomic percent change and absolute measurements were then compared and correlated with associated boney deformities and the Paley classification. RESULTS: The distance from the femoral NV bundle to the ASIS, AIIS, and LT was significantly different compared with the unaffected side. The AIIS absolute distance and AIIS percent change significantly correlated with the neck-shaft angle of the proximal femur. CONCLUSIONS: In patients with CFD, the femoral NV bundle seems to be further from the LT and closer to the AIIS on the affected side when compared with the unaffected side. magnetic resonance imaging may be helpful to understand the course of the femoral NV bundle before reconstruction in patients with CFD; however, the authors recommend identification of the femoral NV bundle before transection of the proximal rectus femoris tendon to provide safe surgical care. LEVEL OF EVIDENCE: Level IV-case-control study of diagnostic studies.

A study of morphological and morphometric variations in Sri Lankan femoral diaphysial nutrient foramina / Estudio de las variaciones morfológicas y morfométricas de los forámenes nutricios diafisarios de fémur en Sri Lanka

SUMMARY: Blood supply to the diaphysis of long bones is majorly through nutrient artery, which enters the bone via nutrient foramen. The present study aims to identify morphological and morphometric variations of nutrient foramina of the femur, which is the largest long bone of the body extending from the pelvis to knee. A sample of 81 Sri Lankan adult femur were analyzed. The mean length of the femur analyzed was 436.93 mm and mean foramen index was 43.52. The majority of the bones had a single nutrient foramen. The predominant location of the nutrient foramen was on the posterior aspect of the bone in the middle third of the shaft according to the study. Majority of foramina were directed distally. The indices on nutrient foramen are important for procedures such as bone grafts and tumor resections, in managing trauma, orthopedic procedures and radiological interpretations.

RESUMEN: El suministro de sangre a la diáfisis de los huesos largos se realiza principalmente a través de la arteria nutricia, que ingresa al hueso a través del foramen nutricio diafisario. El presente estudio tiene como objetivo identificar las variaciones morfológicas y morfométricas del foramen nutricio del fémur, el hueso largo más grande del cuerpo que se extiende desde la pelvis hasta la rodilla. Se analizó una muestra de 81 fémures adultos de individuos de Sri Lanka. La longitud media de los fémures analizados fue de 436,93 mm y el índice de foramen medio fue de 43,52. La mayoría de los huesos tenían un solo foramen nutricio. Según el estudio, la ubicación predominante del foramen nutricio estaba en la cara posterior del hueso en el tercio medio del eje. La mayoría de los forámenes estaban dirigidos distalmente. Los índices sobre el foramen nutricio son importantes para procedimientos tales como injertos óseos y resecciones tumorales, en el manejo de traumatismos, procedimientos ortopédicos e interpretaciones radiológicas.

BONE PERFUSION AND ADIPOSITY BEYOND THE NECROTIC ZONE IN FEMORAL HEAD OSTEONECROSIS: A QUANTITATIVE MRI STUDY.

PURPOSE: To describe bone perfusion and adiposity beyond the necrotic zone with quantitative MRI techniques in osteonecrosis of the femoral head (ONFH). METHOD: In this cross-sectional multicentre study, we recruited patients suffering from late-stage ONFH or hip osteoarthritis. Hip MRI included quantitative MRI sequences: chemical-shift imaging and dynamic contrast-enhanced MRI. We drew regions of interest inside the necrotic zone (inner necrosis and its border) and outside (femoral head, neck and greater trochanter) in ONFH. In the control group, regions of interest were drawn in the femoral head, femoral neck and the greater trochanter. For each region of interest, we measured fat fraction, and calculated semi-quantitative (area under the curve, initial slope) and pharmacokinetic perfusion parameters (Ktrans and Kep). RESULTS: Thirty-two male adults (mean age 58 ±â€¯9 years, range 38-74 years) were included. Sixteen patients formed the ONFH group and fifteen the control group; one was excluded. In the normal-appearing non-necrotic part of the femoral head, fat fraction was not significantly different in comparison with controls (p = 1), but Ktrans was significantly lower than in controls (0.012 ±â€¯0.018 vs. 0.027 ±â€¯0.045; p = 0.05). This perfusion parameter reflects exchanges between blood microvessels and bone marrow. CONCLUSIONS: Our results question the concept of adipose toxicity on the macroscopic scale, and bring up the concept of regional ischemic penumbra that goes beyond the visible necrotic zone. Further studies are required to test these hypotheses in larger populations and earlier disease states.

Medial Femoral Trochlea Osteochondral Flap: Applications for Scaphoid and Lunate Reconstruction.

Vascularized osteochondral flaps are a new technique described for the reconstruction of challenging articular defects of the carpus. The medial femoral trochlea osteochondral flap is supplied by the descending geniculate artery. This osteochondral flap has shown promise in the treatment of recalcitrant scaphoid proximal pole nonunions and advanced avascular necrosis of the lunate. The anatomy, surgical technique, and results are discussed, with clinical cases provided.

Cement arteriovenogram after hip arthroplasty. / Arteriovenograma de cemento óseo en artroplastia de cadera.

Modern cement implantation techniques during hip arthroplasty rely on high intramedullary pressures which can result in cement extrusion towards femoral nutrient vessels, and thus, the occurrence of a particular image in postoperative radiographs (bone cement arterio-venogram). We report a case series of 14 patients in whom a bone cement arterio-venogram was observed after undergoing a cemented hip arthroplasty. No local or systemic complications developed after cementing nor during a mean follow-up of three years. Bone cement arterio-venogram is a radiologic sign that indicates a good cement pressurisation during surgery and is not associated to medical complications or periprosthetic femoral fractures.

Inflammatory Skin-Derived Cytokines Accelerate Osteoporosis in Mice with Persistent Skin Inflammation.

Secondary osteoporosis can also be caused by chronic inflammatory skin disease as well as rheumatoid arthritis or inflammatory bowel disease. However, the exact role of osteoporosis in inflammatory skin conditions has not been elucidated. Using a mouse model of dermatitis, we investigated the pathophysiology of osteoporosis in inflammatory skin conditions and the therapeutic impact of osteoporosis medication on inflammatory skin disease. We employed model mice of spontaneous skin inflammation, specifically overexpressing human caspase-1 in the epidermis. Bone density and the expression of various mRNAs in the femur were examined by micro CT and RT-PCR. The effects of minodronate and anti-RANKL antibody on bone structure, histology, and femur blood flow were studied. The mouse model of skin inflammation showed a marked decrease in bone density compared to wild-type littermates with abnormalities in both bone resorption and formation. Minodronate improved bone density by decreasing osteoclasts, but anti-RANKL antibody did not improve. In the dermatitis model, the blood flow in the bone marrow was decreased, and minodronate restored this parameter. A model of persistent dermatitis exhibited marked osteoporosis, but the impact of chronic dermatitis on osteoporosis has not been thoroughly investigated. We should explore the pathogenesis of osteoporosis in skin inflammatory diseases.

Short-term intermittent parathyroid hormone (1-34) administration increased angiogenesis and matrix metalloproteinase 9 in femora of mature and middle-aged C57BL/6 mice.

NEW FINDINGS: What is the central question of this study? We sought to assess the effects of intermittent parathyroid hormone (1-34) administration on bone angiogenesis, the redistribution of bone marrow blood vessels, and matrix metalloproteinase 9 as a function of advancing age in mice. What is the main finding and its importance? Short-term (i.e. 10 days) intermittent parathyroid hormone (1-34) administration increased the number of small (≤29-µm-diameter) bone marrow blood vessels and augmented matrix metalloproteinase 9. These changes occurred before alterations in trabecular bone. Given the rapid response in bone angiogenesis, this investigation highlights the impact of intermittent parathyroid hormone (1-34) administration on the bone vascular network. ABSTRACT: Intermittent parathyroid hormone (PTH) administration augments bone, stimulates the production of matrix metalloproteinase 9 (Mmp9) and relocates bone marrow blood vessels closer to osteoid seams. Discrepancies exist, however, regarding bone angiogenesis. Given that Mmp9 participates in cellular homing and migration, it might aid in blood vessel relocation. We examined the influence of short-term intermittent PTH administration on angiogenesis, Mmp9 secretion and the distance between blood vessels and bone. Mature (6- to 8-month-old) and middle-aged (10- to 12-month-old) male and female C57BL/6 mice were divided into three groups: control (CON), and 5 (5dPTH) and 10 days (10dPTH) of intermittent PTH administration. Mice were given PBS (50 µl day-1 ) or PTH(1-34) (43 µg kg-1  day-1 ). Frontal sections (5 µm thick) of the right distal femoral metaphysis were triple-immunolabelled to identify endothelial cells (anti-CD31), vascular smooth muscle cells (anti-αSMA) and Mmp9 (anti-Mmp9). Vascular density, Mmp9 density, area and localization, and blood vessel distance from bone were analysed. Blood vessels were analysed according to diameter: 1-29, 30-100 and 101-200 µm. Trabecular bone microarchitecture and bone static and dynamic properties were assessed. No main effects of age were observed for any variable. The density of CD31-labelled blood vessels 1-29 and 30-100 µm in diameter was higher (P < 0.05) and tended (P = 0.055) to be higher, respectively, in 10dPTH versus 5dPTH and CON. Mmp9 was augmented (P < 0.05) in 10dPTH versus the other groups. Mmp9 was closer (P < 0.05) to blood vessels 1-29 µm in diameter and furthest (P < 0.05) from bone. In conclusion, bone angiogenesis occurred by day 10 of intermittent PTH administration, coinciding with augmented Mmp9 secretion near the smallest blood vessels (1-29 µm in diameter).

The structure of cortical bone as revealed by the application of methods for the calcified matrix study.

Calcination and decalcification are basic procedures useful to a morphological approach of a biological, composite material like cortical bone. The study was carried out on a whole human femur conserved in liquid (from an educational collection). Cortical fracturing and SEM observation of vascular canals surface collagen texture was used to study bone deproteination at scalar temperatures (400-1,200°C) and acid bone decalcification at crescent time intervals. Heating burned and vaporized the organic matrix with shrinkage of the bone specimens as documented by the weight loss and transverse surface morphometry. SEM showed a pattern of aligned spherulites at 400°C which maintained the collagen fibrils layout (like a mineral cast), followed by a spherulites fusion progression with the temperature increments. At 1200°C a crystalline-like structure of tightly-packed trapezohendron units. XRD analysis supported the SEM morphology displaying the complete Debey rings of hydroxyapatite and spotted Debey rings of withlockite. Surface Ca and P elution was documented after 12 hr of exposition to the acid solution by dissolution of spherulites and the whole canal surface decalcified in depth after 15 days by SEM-EDAX analysis. The periodic pattern of collagen fibrils was still evident up to 15 days of decalcification together with fine granular deposits of a not-collagenic proteic material, while after 30 days no period was observed in the decalcified fibrils. Collagen mineral cast at 400°C calcination. Complete crystalline transformation at 1200°C. Up to 15 days of decalcification fibrils period maintained.