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Osteoclastogenic potential of peripheral blood mononuclear cells in cleidocranial dysplasia.

Faienza MF, Ventura A, Piacente L, Ciccarelli M, Gigante M, Gesualdo L, Colucci S, Cavallo L, Grano M, Brunetti G - Int J Med Sci (2014)

Bottom Line: The results of this study may help to understand whether in this disease is present an alteration in the bone-resorptive cells, the osteoclasts (OCs).This is in accordance with results arising from flow cytometry experiments demonstrating an high percentage of circulating CD4(+)CD28(+) and CD4(+)CD27(+) T cells, not able to produce osteoclastogenic cytokines.In conclusions, our findings suggest that the heterozygous deletion of RUNX2 in this CCD patient did not alter the osteoclastogenic potential of PBMCs in vitro.

View Article: PubMed Central - PubMed

Affiliation: 1. Department of Biomedical Sciences and Human Oncology, Section of Pediatrics, University of Bari, Bari, Italy;

ABSTRACT
Cleidocranial dysplasia (CCD) is an autosomal dominant skeletal dysplasia characterized by hypoplastic or aplastic clavicles, dental abnormalities, and delayed closure of the cranial sutures. In addition, mid-face hypoplasia, short stature, skeletal anomalies and osteoporosis are common. We aimed to evaluate osteoclastogenesis in a child (4 years old), who presented with clinical signs of CCD and who have been diagnosed as affected by deletion of RUNX2, master gene in osteoblast differentiation, but also affecting T cell development and indirectly osteoclastogenesis. The results of this study may help to understand whether in this disease is present an alteration in the bone-resorptive cells, the osteoclasts (OCs). Unfractionated and T cell-depleted Peripheral Blood Mononuclear Cells (PBMCs) from patient were cultured in presence/absence of recombinant human M-CSF and RANKL. At the end of the culture period, OCs only developed following the addition of M-CSF and RANKL. Moreover, real-time PCR experiment showed that freshly isolated T cells expressed the osteoclastogenic cytokines (RANKL and TNFα) at very low level, as in controls. This is in accordance with results arising from flow cytometry experiments demonstrating an high percentage of circulating CD4(+)CD28(+) and CD4(+)CD27(+) T cells, not able to produce osteoclastogenic cytokines. Also RANKL, OPG and CTX serum levels in CCD patient are similar to controls, whereas QUS measurements showed an osteoporotic status (BTT-Z score -3.09) in the patient. In conclusions, our findings suggest that the heterozygous deletion of RUNX2 in this CCD patient did not alter the osteoclastogenic potential of PBMCs in vitro.

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Related in: MedlinePlus

CD14+/CD16+ and CD4+/CD25high cells in CCD patient. Flow cytometry analysis of CD14+/CD16+ and CD4+/CD25high cells in peripheral blood cells revealed that CD14+/CD16+ was significantly higher in patient (B) compared with the age-matched normal controls (A), whereas no significant differences were detected in CD4+/CD25high cells among patient (D) and controls (C).
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Figure 2: CD14+/CD16+ and CD4+/CD25high cells in CCD patient. Flow cytometry analysis of CD14+/CD16+ and CD4+/CD25high cells in peripheral blood cells revealed that CD14+/CD16+ was significantly higher in patient (B) compared with the age-matched normal controls (A), whereas no significant differences were detected in CD4+/CD25high cells among patient (D) and controls (C).

Mentions: Peripheral blood cells from CCD patient and controls were subjected to flow cytometry analysis to evaluate the immunophenotype of monocytes and T cells. In particular, in the patient we found a percentage (1.7 ± 0.5%) of CD14+/CD16+ cells, which can be considered committed OC precursors 31, that are absent in controls (P < 0.01), Figure 2A-2B. Moreover, we characterized both CD4+ and CD8+ T cells focusing on markers of T-cell activation, proliferation, and differentiation. No significant differences for CD4+CD25high population were observed between patient and controls (Figure 2C-2D). Staining of freshly peripheral blood T cells revealed that costimulatory molecules (CD28, CD27, CD62L, CD69) were highly expressed on both CD4+ and CD8+ T cells cells from CCD patient (Figure 3-4). Moreover, we also showed that in CCD patient the subset CD4+/CD28- is smaller than controls (Figure 3). This is an important finding because this subset is involved in the production of TNFα, and consequently supporting osteoclastogenesis. Similarly, we also found that the subset CD8+/CD27- include a small percentage of cells in patient (Figure 4). This is relevant because this subset is involved in the production of inflammatory cytokines.


Osteoclastogenic potential of peripheral blood mononuclear cells in cleidocranial dysplasia.

Faienza MF, Ventura A, Piacente L, Ciccarelli M, Gigante M, Gesualdo L, Colucci S, Cavallo L, Grano M, Brunetti G - Int J Med Sci (2014)

CD14+/CD16+ and CD4+/CD25high cells in CCD patient. Flow cytometry analysis of CD14+/CD16+ and CD4+/CD25high cells in peripheral blood cells revealed that CD14+/CD16+ was significantly higher in patient (B) compared with the age-matched normal controls (A), whereas no significant differences were detected in CD4+/CD25high cells among patient (D) and controls (C).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3936030&req=5

Figure 2: CD14+/CD16+ and CD4+/CD25high cells in CCD patient. Flow cytometry analysis of CD14+/CD16+ and CD4+/CD25high cells in peripheral blood cells revealed that CD14+/CD16+ was significantly higher in patient (B) compared with the age-matched normal controls (A), whereas no significant differences were detected in CD4+/CD25high cells among patient (D) and controls (C).
Mentions: Peripheral blood cells from CCD patient and controls were subjected to flow cytometry analysis to evaluate the immunophenotype of monocytes and T cells. In particular, in the patient we found a percentage (1.7 ± 0.5%) of CD14+/CD16+ cells, which can be considered committed OC precursors 31, that are absent in controls (P < 0.01), Figure 2A-2B. Moreover, we characterized both CD4+ and CD8+ T cells focusing on markers of T-cell activation, proliferation, and differentiation. No significant differences for CD4+CD25high population were observed between patient and controls (Figure 2C-2D). Staining of freshly peripheral blood T cells revealed that costimulatory molecules (CD28, CD27, CD62L, CD69) were highly expressed on both CD4+ and CD8+ T cells cells from CCD patient (Figure 3-4). Moreover, we also showed that in CCD patient the subset CD4+/CD28- is smaller than controls (Figure 3). This is an important finding because this subset is involved in the production of TNFα, and consequently supporting osteoclastogenesis. Similarly, we also found that the subset CD8+/CD27- include a small percentage of cells in patient (Figure 4). This is relevant because this subset is involved in the production of inflammatory cytokines.

Bottom Line: The results of this study may help to understand whether in this disease is present an alteration in the bone-resorptive cells, the osteoclasts (OCs).This is in accordance with results arising from flow cytometry experiments demonstrating an high percentage of circulating CD4(+)CD28(+) and CD4(+)CD27(+) T cells, not able to produce osteoclastogenic cytokines.In conclusions, our findings suggest that the heterozygous deletion of RUNX2 in this CCD patient did not alter the osteoclastogenic potential of PBMCs in vitro.

View Article: PubMed Central - PubMed

Affiliation: 1. Department of Biomedical Sciences and Human Oncology, Section of Pediatrics, University of Bari, Bari, Italy;

ABSTRACT
Cleidocranial dysplasia (CCD) is an autosomal dominant skeletal dysplasia characterized by hypoplastic or aplastic clavicles, dental abnormalities, and delayed closure of the cranial sutures. In addition, mid-face hypoplasia, short stature, skeletal anomalies and osteoporosis are common. We aimed to evaluate osteoclastogenesis in a child (4 years old), who presented with clinical signs of CCD and who have been diagnosed as affected by deletion of RUNX2, master gene in osteoblast differentiation, but also affecting T cell development and indirectly osteoclastogenesis. The results of this study may help to understand whether in this disease is present an alteration in the bone-resorptive cells, the osteoclasts (OCs). Unfractionated and T cell-depleted Peripheral Blood Mononuclear Cells (PBMCs) from patient were cultured in presence/absence of recombinant human M-CSF and RANKL. At the end of the culture period, OCs only developed following the addition of M-CSF and RANKL. Moreover, real-time PCR experiment showed that freshly isolated T cells expressed the osteoclastogenic cytokines (RANKL and TNFα) at very low level, as in controls. This is in accordance with results arising from flow cytometry experiments demonstrating an high percentage of circulating CD4(+)CD28(+) and CD4(+)CD27(+) T cells, not able to produce osteoclastogenic cytokines. Also RANKL, OPG and CTX serum levels in CCD patient are similar to controls, whereas QUS measurements showed an osteoporotic status (BTT-Z score -3.09) in the patient. In conclusions, our findings suggest that the heterozygous deletion of RUNX2 in this CCD patient did not alter the osteoclastogenic potential of PBMCs in vitro.

Show MeSH
Related in: MedlinePlus