Mobilization of CD34+-progenitor cells in patients with severe trauma.

Circulating CD34+ progenitor cells () gained importance in the field of regenerative medicine due to their potential to home in on injury sites and differentiate into cells of both endothelial and osteogenic lineages. In this study, we analyzed the mobilization kinetics and the numbers of CD34+, CD31+, CD45+, and CD133+ cells in twenty polytrauma patients (n = 13 male, n = 7 female, mean age 46.5±17.2 years, mean injury severity score (ISS) 35.8±12.5 points). In addition, the endothelial differentiation capacity of enriched CD34+cells was assessed by analyzing DiI-ac-LDL/lectin uptake, the expression of endothelial markers, and the morphological characteristics of these cells in Matrigel and spheroid cultures. We found that on days 1, 3, and 7 after a major trauma, the number of CD34+cells increased from 6- up to 12-fold (p<0.0001) over the number of CD34+cells from a control population of healthy, age-matched volunteers. The numbers of CD31+ cells were consistently higher on days 1 (1.4-fold, p<0.01) and 7 (1.3-fold, p<0.01), whereas the numbers of CD133+ cell did not change during the time course of investigation. Expression of endothelial marker molecules in CD34+cells was significantly induced in the polytrauma patients. In addition, we show that the CD34+ cell levels in severely injured patients were not correlated with clinical parameters, such as the ISS score, the acute physiology and chronic health evaluation II score (APACHE II), as well as the sequential organ failure assessment score (SOFA-2). Our results clearly indicate that pro-angiogenic cells are systemically mobilized after polytrauma and that their numbers are sufficient for the development of novel therapeutic models in regenerative medicine

Mobilization of CD34+-progenitor cells in patients with severe trauma.

Circulating CD34+ progenitor cells () gained importance in the field of regenerative medicine due to their potential to home in on injury sites and differentiate into cells of both endothelial and osteogenic lineages. In this study, we analyzed the mobilization kinetics and the numbers of CD34+, CD31+, CD45+, and CD133+ cells in twenty polytrauma patients (n = 13 male, n = 7 female, mean age 46.5±17.2 years, mean injury severity score (ISS) 35.8±12.5 points). In addition, the endothelial differentiation capacity of enriched CD34+cells was assessed by analyzing DiI-ac-LDL/lectin uptake, the expression of endothelial markers, and the morphological characteristics of these cells in Matrigel and spheroid cultures. We found that on days 1, 3, and 7 after a major trauma, the number of CD34+cells increased from 6- up to 12-fold (p<0.0001) over the number of CD34+cells from a control population of healthy, age-matched volunteers. The numbers of CD31+ cells were consistently higher on days 1 (1.4-fold, p<0.01) and 7 (1.3-fold, p<0.01), whereas the numbers of CD133+ cell did not change during the time course of investigation. Expression of endothelial marker molecules in CD34+cells was significantly induced in the polytrauma patients. In addition, we show that the CD34+ cell levels in severely injured patients were not correlated with clinical parameters, such as the ISS score, the acute physiology and chronic health evaluation II score (APACHE II), as well as the sequential organ failure assessment score (SOFA-2). Our results clearly indicate that pro-angiogenic cells are systemically mobilized after polytrauma and that their numbers are sufficient for the development of novel therapeutic models in regenerative medicine

Mobilization of CD34+-progenitor cells in patients with severe trauma

Circulating CD34+ progenitor cells () gained importance in the field of regenerative medicine due to their potential to home in on injury sites and differentiate into cells of both endothelial and osteogenic lineages. In this study, we analyzed the mobilization kinetics and the numbers of CD34+, CD31+, CD45+, and CD133+ cells in twenty polytrauma patients (n = 13 male, n = 7 female, mean age 46.5±17.2 years, mean injury severity score (ISS) 35.8±12.5 points). In addition, the endothelial differentiation capacity of enriched CD34+cells was assessed by analyzing DiI-ac-LDL/lectin uptake, the expression of endothelial markers, and the morphological characteristics of these cells in Matrigel and spheroid cultures. We found that on days 1, 3, and 7 after a major trauma, the number of CD34+cells increased from 6- up to 12-fold (p<0.0001) over the number of CD34+cells from a control population of healthy, age-matched volunteers. The numbers of CD31+ cells were consistently higher on days 1 (1.4-fold, p<0.01) and 7 (1.3-fold, p<0.01), whereas the numbers of CD133+ cell did not change during the time course of investigation. Expression of endothelial marker molecules in CD34+cells was significantly induced in the polytrauma patients. In addition, we show that the CD34+ cell levels in severely injured patients were not correlated with clinical parameters, such as the ISS score, the acute physiology and chronic health evaluation II score (APACHE II), as well as the sequential organ failure assessment score (SOFA-2). Our results clearly indicate that pro-angiogenic cells are systemically mobilized after polytrauma and that their numbers are sufficient for the development of novel therapeutic models in regenerative medicine

Distribution of measurement values in the control group (Contr.) and the group with severe trauma (ST) at days 1 (d1), 3 (d3), and 7 (d7).

<p>The relative numbers (%) of CD34⁺ (d1, d3, and d7), CD31 (d1 and 7), and CD34⁺/CD45⁻ cells (d 7) were significantly increased in the ST group during the period of investigation, as compared with the control group (Mann-Whitney-U-test, see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0097369#pone-0097369-t003" target="_blank">Table 3</a> for p-values). In contrast, the number of CD45⁺ cells was significantly lower on day seven in the ST group. The total number of CD34⁺ cells (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0097369#pone-0097369-t003" target="_blank">Table 3</a>) was sufficient for isolation and further culturing procedures.</p

Pearson's correlation analysis.

<p>Pearson's correlation analysis.</p

LDL-lectin and immunofluorescence staining analyses of CD34+-cells a-b: DiI-ac-LDL/lectin double staining of CD34⁺ cell cultures.

<p><b>a</b>: control group, <b>b</b>: ST group. <b>c</b>: phase contrast/Hoechst 33258 staining of purified CD34+ST cultures (cell nuclei stained in blue), same picture as <b>d</b>: CD31 (red)/CD34 (green)/Hoechst 33258 (blue) – immunofluorescence staining showing positive reaction with cultured cells. <b>e-f</b>: CD146 (green)/Hoechst 33258 (blue) – immunofluorescence staining showing low cell numbers with positive staining in the control group (e) but positive reactions in cultured cells isolated from patients with severe trauma (f). All scale bars are 100 µm.</p

Polytrauma patients recruited to the study.

<p>Polytrauma patients recruited to the study.</p

Immunohistological stainings a–d: Morphology of cells cultured on Matrigel, a: control group, b: ST group, c: CD34-depleted cell fraction (negative control), d: HUVEC cells (positive control).

<p>No formation of cord-like structures could be detected in control- and CD34-depleted cell cultures. Formation of cord-like structures was present in the ST group, but to a much lower extent as compared with HUVECs. <b>e–g</b>: Endothelial cell sprouting from carboxymethylcellulose spheroids. No formation of sprouts could be detected in the control group (<b>e</b>) whereas primitive endothelial sprouts were monitored in the ST-group (<b>f</b>). However, these formations were much less compared to HUVEC-sprout formation (<b>g</b>). All scale bars are 100 µm.</p