Systemic Regulation of Bone Marrow Stromal Cytokines After Severe Trauma.

BACKGROUND: Traumatic injury generates a prolonged hypercatecholamine state that is associated with reduced growth of bone marrow erythroid progenitors mediated by the bone marrow stroma. The bone marrow stroma is made up of many cells including fibroblasts, which respond to inflammatory stimuli and alter the cytokine profile. We hypothesized that trauma plasma would increase bone marrow stromal fibroblast expression of interleukin-6 (IL-6), granulocyte colony-stimulating factor (G-CSF), erythropoietin (EPO), stem cell factor (SCF), and activation of nuclear factor kappa-light-chain-enhancer of activated B cells and correlate with injury severity and anemia. MATERIALS AND METHODS: Plasma from 15 trauma patients was cultured with bone marrow fibroblast cells and compared with that from healthy volunteers. At 6, 24, and 48 h, the expression of IL-6, G-CSF, EPO, SCF, and the activation of nuclear factor kappa-light-chain-enhancer of activated B cells were measured using quantitative polymerase chain reaction. The influence of trauma plasma on cytokine expression was further stratified by injury severity score (ISS). RESULTS: The average hemoglobin significantly decreased from admission to discharge (10.7 ± 2.5 to 9.2 ± 1.1 g/dL, P < 0.04). The discharge hemoglobin significantly decreased by 14% from the admission hemoglobin. After 48 h, trauma plasma significantly increased IL-6, G-CSF, and EPO bone marrow fibroblast expression when compared with normal plasma. When stratified by ISS, IL-6, G-CSF, and EPO, bone marrow fibroblast expression was highest in the trauma plasma ISS 27-41 group and was significantly elevated compared with normal plasma. When SCF expression was stratified by ISS, there was a significant increase in expression in ISS 27-41. Higher ISS was also associated with a larger decrease in hemoglobin despite no difference in total blood transfusions. CONCLUSIONS: Severe trauma can systemically increase IL-6, G-CSF, and EPO expression in bone marrow stroma. Increased hematopoietic cytokine expression after traumatic injury correlated with a hypercatecholamine state, anemia, and injury severity.

The Postinjury Inflammatory State and the Bone Marrow Response to Anemia.

RATIONALE: The pathophysiology of persistent injury-associated anemia is incompletely understood, and human data are sparse. OBJECTIVES: To characterize persistent injury-associated anemia among critically ill trauma patients with the hypothesis that severe trauma would be associated with neuroendocrine activation, erythropoietin dysfunction, iron dysregulation, and decreased erythropoiesis. METHODS: A translational prospective observational cohort study comparing severely injured, blunt trauma patients who had operative fixation of a hip or femur fracture (n = 17) with elective hip repair patients (n = 22). Bone marrow and plasma obtained at the index operation were assessed for circulating catecholamines, systemic inflammation, erythropoietin, iron trafficking pathways, and erythroid progenitor growth. Bone marrow was also obtained from healthy donors from a commercial source (n = 8). MEASUREMENTS AND MAIN RESULTS: During admission, trauma patients had a median of 625 ml operative blood loss and 5 units of red blood cell transfusions, and Hb decreased from 10.5 to 9.3 g/dl. Compared with hip repair, trauma patients had higher median plasma norepinephrine (21.9 vs. 8.9 ng/ml) and hepcidin (56.3 vs. 12.2 ng/ml) concentrations (both P < 0.05). Bone marrow erythropoietin and erythropoietin receptor expression were significantly increased among patients undergoing hip repair (23% and 14% increases, respectively; both P < 0.05), but not in trauma patients (3% and 5% increases, respectively), compared with healthy control subjects. Trauma patients had lower bone marrow transferrin receptor expression than did hip repair patients (57% decrease; P < 0.05). Erythroid progenitor growth was decreased in trauma patients (39.0 colonies per plate; P < 0.05) compared with those with hip repair (57.0 colonies per plate; P < 0.05 compared with healthy control subjects) and healthy control subjects (66.5 colonies per plate). CONCLUSIONS: Severe blunt trauma was associated with neuroendocrine activation, erythropoietin dysfunction, iron dysregulation, erythroid progenitor growth suppression, and persistent injury-associated anemia. Clinical trial registered with www.clinicaltrials.gov (NCT 02577731).

Chronic stress induces persistent low-grade inflammation.

INTRODUCTION: This study sought to determine if the systemic cytokine profile of rodents subjected to chronic restraint stress leads to persistent low-grade inflammation. METHODS: Male Sprague-Dawley rats were subjected to restraint stress for a total of seven or fourteen days. Urine norepinephrine (NE), plasma interleukin 6 (IL-6), tumor necrosis factor alpha (TNF-α), C-reactive protein (CRP) were assessed with ELISA. Liver expression of IL-6 and TNF-α were assessed with real time PCR. RESULTS: Chronic stress at 7 and 14 days sequentially increased plasma acute phase reactants (NE, IL-6, TNF-α, and CRP), liver IL-6 expression, hematopoietic progenitor cell mobilization, and decreased erythroid progenitor colony growth. Weight gain was reduced by chronic stress compared to each models' naïve counterpart. CONCLUSIONS: Combining this model with trauma and sepsis models will allow evaluation of the contribution of persistent inflammation in disease progression and outcomes.

Persistent injury-associated anemia in aged rats.

BACKGROUND: Hypercatecholaminemia and bone marrow dysfunction have been implicated in the pathophysiology of persistent-injury associated anemia. The elderly may be vulnerable to this phenomenon due to high basal and peak catecholamine levels, impaired erythroid progenitor growth, and baseline anemia. We hypothesized that aged F344-BN rats subjected to severe trauma and chronic stress would have persistent injury-associated anemia. METHODS: Male F344-BN rats age 25months were randomly allocated to: naïve (n=8), lung contusion (LC, n=9), LC followed by daily chronic restraint stress (LC/CS, n=9), LC followed immediately by hemorrhagic shock (LCHS, n=8), and LCHS followed by daily CS (LCHS/CS, n=8). Urine norepinephrine was measured on days one and seven. Locomotor testing was performed on day five. Bone marrow cellularity, hematopoietic progenitor growth, and peripheral blood hemoglobin levels were assessed at sacrifice on day seven. Data are presented as mean±standard deviation, *p<0.05 vs. naïve. RESULTS: Norepinephrine levels (ng/mL) were significantly elevated one day after LCHS (420±239* vs. naïve: 97±71) and LCHS/CS (375±185*), and remained significantly elevated on day seven for LCHS/CS (359±99*), but not LCHS (212±130). On locomotor testing, groups subjected to CS traveled shorter distances at lower velocities and spent less time in the center of the cage. Colony forming units-erythroid (colonies/plate), representing late erythroid progenitors, were significantly decreased after LC/CS (40±1* vs. naïve: 47±4), LCHS (40±1*), and LCHS/CS (38±3*). LCHS/CS animals had significantly lower hemoglobin (g/dL) than naïve animals (13.3±1.3* vs. naïve: 15.2±0.9). CONCLUSIONS: Persistent injury-associated anemia occurs in aged rats. Further research is needed to determine whether the pathophysiology of this phenomenon differs from that of younger rats, and to translate these findings to elderly trauma patients.

Persistent injury-associated anemia and aging: Novel insights.

BACKGROUND: Hypercatecholaminemia and bone marrow dysfunction have been implicated in the pathophysiology of persistent injury-associated anemia. The elderly may be more vulnerable to bone marrow dysfunction due to high basal and peak catecholamine levels and impaired hematopoietic progenitor growth. We hypothesized that aging would adversely affect persistent injury-associated anemia. METHODS: Male Sprague-Dawley rats aged 8 to 9 weeks and F344-BN rats aged 25 months were randomized to naive controls, lung contusion plus hemorrhagic shock (LCHS), and LCHS plus daily chronic restraint stress (LCHS/CS). Urine norepinephrine was measured on Days 1 and 7. Mobilization of hematopoietic progenitor cells (HPCs), bone marrow colony-forming units-erythroid growth, and peripheral blood hemoglobin, mean corpuscular volume (MCV), and red cell distribution width (RDW) were assessed on Day 7 (*p < 0.05 young vs. aged counterpart by one-way analysis of variance). RESULTS: Aged rats had higher norepinephrine levels at naive baseline (97* vs. 27 ng/mL) and 7 days following LCHS/CS when compared with young (359* vs. 127 ng/mL). Following LCHS/CS, HPC mobilization was greater among young rats when compared with aged (5.4 vs. 2.5%). Colony-forming units-erythroid growth was lower among aged animals for each group (naive: 47* vs. 65; LCHS: 40* vs. 50; LCHS/CS: 38* vs. 44 cells/plate). Aged naive rats had higher initial hemoglobin (15.2* vs. 14.3 g/dL) but lower MCV (48* vs. 59 fL/cell) and larger RDW at baseline and greater differences 7 days after LCHS/CS (MCV: 46* vs. 60 fL/cell; RDW: 17.4* vs. 16.3%). CONCLUSIONS: Compared with young rats, aged rats had less HPC mobilization despite elevated basal and peak norepinephrine. Aged rats were disproportionately affected by impaired hematopoietic progenitor growth and an iron-restricted red blood cell phenotype at baseline, which persisted 7 days after injury. Further research is needed to assess how the clinical approach to persistent injury-associated anemia should differ for elderly trauma patients.

Clonidine restores vascular endothelial growth factor expression and improves tissue repair following severe trauma.

BACKGROUND: We hypothesized that clonidine and propranolol would increase VEGF and VEGF-receptor expression and promote lung healing following severe trauma and chronic stress. METHODS: Sprague-Dawley rats were subjected to lung contusion (LC), lung contusion/hemorrhagic shock (LCHS), or lung contusion/hemorrhagic shock/daily restraint stress (LCHS/CS). Clonidine and propranolol were administered daily. On day seven, lung VEGF, VEGFR-1, VEGFR-2, and HMGB1 were assessed by PCR. Lung injury was assessed by light microscopy (*p < 0.05). RESULTS: Clonidine increased VEGF expression following LCHS (43%*) and LCHS/CS (46%*). Clonidine increased VEGFR-1 and R-2 expression following LCHS/CS (203%* and 47%*, respectively). Clonidine decreased HMGB1 and TNF-alpha expression following LCHS/CS (22%* and 58%*, respectively.) Clonidine decreased inflammatory cell infiltration and total Lung Injury Score following LCHS/CS. Propranolol minimally affected VEGF and did not improve lung healing. CONCLUSIONS: Clonidine increased VEGF and VEGF-receptor expression, decreased HMGB1 expression, decreased lung inflammation, and improved lung tissue repair.