Petra Algenstaedt

Neural driven angiogenesis by overexpression of nerve growth factor.

Mechanisms regulating angiogenesis are crucial in adjusting tissue perfusion on metabolic demands. We demonstrate that overexpression of nerve growth factor (NGF) in brown adipose tissue (BAT) of NGF-transgenic mice elevates both mRNA and protein levels of vascular endothelial growth factor (VEGF) and VEGF-receptors. Increased vascular permeability, leukocyte–endothelial interactions (LEI), and tissue perfusion were measured using intravital microscopy. NGF-stimulation of adipocytes and endothelial cells elevates mRNA expression of VEGF and its receptors, an effect blocked by NGF neutralizing antibodies. These data suggest an activation of angiogenesis as a result of both: stimulation of adipozytes and direct mitogenic effects on endothelial cells. The increased nerve density associated with vessels strengthened our hypothesis that tissue perfusion is regulated by neural control of vessels and that the interaction between the NGF and VEGF systems is the critical driver for the activated angiogenic process. The interaction of VEGF- and NGF-systems gives new insights into neural control of organ vascularization and perfusion.

Sequential changes in vessel formation and micro-vascular function during bone repair

Background Angiogenesis, the process of new vessel formation from a pre-existing vascular network, is essential for bone development and repair. New vessel formation and microvascular functions are crucial during bone repair, not only for sufficient nutrient supply, transport of macromolecules and invading cells, but also because they govern the metabolic microenvironment. Despite its central role, very little is known about the initial processes of vessel formation and microvascular function during bone repair. Methods To visualize and quantify the process of vessel formation and microvascular function during bone repair, we transplanted neonatal femora with a substantial defect into dorsal skin-fold chambers in severe combined immunodeficient (SCID) mice for continuous noninvasive in-vivo evaluation. We employed intravital microscopic techniques to monitor effective microvascular permeability, functional vascular density, blood flow rate and leukocyte flux repeatedly over 16 days. Oxytetracyclin and v. Kossa/v. Giesson staining was performed to quantify the calcification process in vivo and in vitro. Results Development of a hematoma surrounding the defect area was the initial event, which was accompanied by a significant increase in microvascular permeability and blood flow rate. With absorption of the hematoma and vessel maturation, permeability decreased continuously, while vascular density and tissue perfusion increased. Histological evaluation revealed that the remodeling of the substantial defect prolonged the in-vivo monitored calcification process. Interpretation The size of the initial substantial defect correlated positively with increased permeability, suggesting improved release of permeability-inducing cytokines. The unchanged permeability in the control group with boiled bones and a substantial defect corroborated these findings. The adaptation to increasing metabolic demands was initially mediated by increased blood flow rate, later with increasing vascular density through increased tissue perfusion rate. These insights into the sequence of microvascular alterations may assist in the development of targeted drug delivery therapies and caution against the use of permeability-altering drugs during bone healing.

Bilaterally increased VEGF-levels in muscles during experimental unilateral callus distraction

Angiogenesis is essential for wound healing and proliferative processes such as bone formation and repair. Since increased expression of the vascular endothelial growth factor (VEGF) stimulates bone formation, it can be hypothesized that surgical procedures leading to a systemic increase of VEGF for instance during wound healing, influence enchondral ossification processes and might be responsible for observed growth phenomena during callus distraction. To study the mechanisms of angiogenesis in soft tissue during unilateral callus distraction, lengthening of the right tibia was performed in 12 beagles. After osteotomy, application of a ring fixator and after five latency days, distraction was started for 25 days. A control group of four additional beagles underwent no surgical procedure. Subsequent to the distraction period (Group A), muscle samples from six beagles were taken from the distracted side (ds) and the contralateral non‐distracted side (n‐ds), six beagles underwent an additional consolidation period of 25 days (Group B). Samples were analyzed for VEGF, VEGFR‐1 and VEGFR‐2 mRNA expression using real‐time PCR and protein expression using Western Blot analysis. Muscles from both extremities showed significantly increased expression of VEGF and its cognate receptors VEGFR‐½. Expression decreased significantly after the consolidation period, whereby the level at the non‐distracted side decreased more than the level at the distracted side. Interestingly VEGF and VEGFR‐1 levels at the non‐distracted side were significantly higher than at the distracted side. In contrast VEGFR‐2, the receptor that mediates endothelial cell proliferation, showed higher levels at the distracted than at the non‐distracted side. These findings indicate that callus distraction results not only in locally increased expression of VEGF and its receptors, but leads also to increased VEGF and VEGFR‐½ levels at distant sides and might therefore be responsible for the observed growth phenomena during callus distraction.

Chemokine profile of disc degeneration with acute or chronic pain

OBJECT Disc-related disorders such as herniation and chronic degenerative disc disease (DDD) are often accompanied by acute or chronic pain. Different mediators have been identified in the development of radicular pain and DDD. Previous studies have not analyzed individual cytokine profiles discriminating between acute sciatic and chronic painful conditions, nor have they distinguished between different anatomical locations within the disc. The aim of this study was to elucidate the protein biochemical mechanisms in DDD. METHODS The authors determined expression levels of matrix metalloproteinase-3, transforming growth factor-β (TGF-β), tumor necrosis factor-α, interleukin-1α, and pro-substance P using enzyme-linked immunosorbent assay and Western blot analyses in patients suffering from DDD (n = 7), acute back pain due to herniated discs with radiculopathy (n = 7), and a control group (n = 7). Disc tissue samples from the anulus fibrosus (AF) and nucleus pulposus (NP) were analyzed. Statistical analysis was performed using nonparametric tests. RESULTS A distinct distribution of cytokines was found in different anatomical regions of intervertebral discs in patients with DDD and herniated NP. Increased TGF-β levels were predominantly found in DDD. Matrix metalloproteinase-3 was increased in acute herniated disc material. Increased levels of substance P were found in patients suffering from DDD but not in patients with disc herniation. The data showed significantly higher levels of proinflammatory cytokines in the AF and NP of patients with DDD, and the expression levels in the AF were even higher than in the NP, suggesting that the inflammatory response initiates from the AF. CONCLUSIONS These results highlight the complex mechanisms involved during disc degeneration and the need to distinguish between acute and chronic processes as well as different anatomical regions, namely the AF and NP. They also highlight potential problems in disc nucleus replacement therapies because the results suggest a biochemical link between AF and NP cytokine expression.

Comparison Between Minimally Invasive Surgery and Conventional Open Surgery for Patients With Spinal Metastasis: A Prospective Propensity Score-matched Study.

Study Design. Prospective propensity score-matched study. Objective. To compare the outcomes of minimal invasive surgery (MIS) and conventional open surgery for spinal metastasis patients. Summary of Background Data. There is lack of knowledge on whether MIS is comparable to conventional open surgery in treating spinal metastasis. Methods. Patients with spinal metastasis requiring surgery from January 2008 to December 2010 in two spine centers were recruited. The demographic, preoperative, operative, perioperative and postoperative data were collected and analyzed. Thirty MIS patients were matched with 30 open surgery patients using propensity score matching technique with a match tolerance of 0.02 based on the covariate age, tumor type, Tokuhashi score, and Tomita score. Results. Both groups had significant improvements in Eastern Cooperative Oncology Group (ECOG), Karnofsky scores, visual analogue scale (VAS) for pain and neurological status postoperatively. However, the difference comparing the MIS and open surgery group was not statistically significant. MIS group had significantly longer instrumented segments (5.5 ± 3.1) compared with open group (3.8 ± 1.7). Open group had significantly longer decompressed segment (1.8 ± 0.8) than MIS group (1.0 ± 1.0). Open group had significantly more blood loss (2062.1 ± 1148.0 mL) compared with MIS group (1156.0 ± 572.3 mL). More patients in the open group (76.7%) needed blood transfusions (with higher average units of blood transfused) compared with MIS group (40.0%). Fluoroscopy time was significantly longer in MIS group (116.1 ± 63.3 s) compared with open group (69.9 ± 42.6 s). Open group required longer hospitalization (21.1 ± 10.8 days) compared with MIS group (11.0 ± 5.0 days). Conclusion. This study demonstrated that MIS resulted in comparable outcome to open surgery for patients with spinal metastasis but has the advantage of less blood loss, blood transfusions, and shorter hospital stay. Level of Evidence: 3

Sunitinib treatment reduces tumor growth and limits changes in microvascular properties after minor surgical intervention in an in vivo model of secondary breast cancer growth in bone.

Surgical interventions can alter the balance between pro‐ and anti‐angiogenic growth factors and thereby modulate tumor growth. Since the microcirculatory properties of tumors underlie organ‐specific differences, the microhemodynamic characteristics of bone metastasis have not yet been fully described. Angiogenesis inhibitors are increasingly being used to treat advanced stages of cancer. We hypothesized that the anti‐angiogenic drug sunitinib abrogates alterations in microvascular properties following a minor surgical intervention in an in vivo model of secondary breast cancer growth in the bone.