Biomedical subjects

Dennis von Heimburg

Publications and source records attributed to Dennis von Heimburg.

13 recordsLinked to original sources

PubMed2006

Human serum from platelet-poor plasma for the culture of primary human preadipocytes.

In adipose tissue engineering, the use of human serum is essential to achieve the goal of an autologous system. Serum from conventional human plasma (SCP) contains platelet-derived growth factor (PDGF), a growth factor known to be both a potent inhibitor of adipose differentiation and also the most important stimulator of proliferation in human serum. Serum from platelet-poor plasma (SPPP) is considered to be PDGF-deprived and should therefore inhibit the differentiation of preadipocytes to adipocytes to a lesser extent. Effective cultivation of preadipocytes with SPPP requires compensating for the missing stimulatory PDGF effect on proliferation. However, the addition of other growth factors to the media needs to provide stimulation of proliferation without significant inhibition of differentiation. Primary human preadipocytes were isolated from adipose tissue samples of 10 healthy human donors and cultured under four different medium conditions (SCP, SPPP, SPPP + 1 nM basic fibroblast growth factor [bFGF], and SPPP + 1 nM epidermal growth factor [EGF]) for five generations. Proliferation activity and differentiation capacity were assessed for each sample, generation, and culture condition by calculating doubling time and measuring glycerol-3-phosphate dehydrogenase (GPDH)-specific activity. The use of SPPP resulted in a marked rise in GPDH activity compared with the cells cultured with SCP. Supplementing SPPP with 1 nM bFGF or EGF increased proliferation activity significantly. SPPP can be considered superior to SCP for the culture of primary human preadipocytes in adipose tissue engineering in terms of proliferation activity and differentiation capacity.

Adipocytes↗

PubMed2006

Biomaterials for adipose tissue engineering.

There is high clinical need for an adequate reconstruction of soft tissue defects as found after tumor resections, deep burns or severe trauma. A promising solution for these defects is adipose tissue engineering, with adult stem cells of the adipose tissue, implanted on 3D biomaterials. These adipogenic precursor cells survive ischemia better than mature adipocytes and have the potency to proliferate and differentiate into fat cells after transplantation. They can be yielded from excised adipose tissue or liposuction material. When preadipocytes are seeded on carriers for the generation of adipose tissue, chemical composition, mechanical stability and 3D architecture of the construct are crucial factors. They ensure cellular penetration into the construct, sufficient proliferation on the material and full differentiation inside the construct after transplantation. In hydrogels, it is especially the use and combination of growth factors that determine the overall outcome of the applied biopolymer. Over recent years, in vivo trials in particular have allowed significant insights into the potential, the perspectives, but also the current difficulties and draw-backs in adipose tissue engineering. This review focuses on the main strategies in adipose tissue regeneration, compares the various materials that have been used as carrier matrices so far and considers them in light of the challenges they have yet to meet.

Adipocytes↗

PubMed2005

Efficacy and safety of polyacrylamide hydrogel for facial soft-tissue augmentation.

BACKGROUND: Polyacrylamide is a nonabsorbable soft-tissue filler that has been used for body contouring for more than 10 years. Aquamid is a soft-tissue filler containing polyacrylamide hydrogel. Safety and aesthetic results after injection of polyacrylamide hydrogel for facial soft-tissue enhancement were assessed at a 1-year follow-up. METHODS: In a prospective, multicenter study, 251 patients were enrolled and assigned for injection of polyacrylamide hydrogel (Aquamid) as a means of facial augmentation and enhancement of facial contours. Subjects with facial contour deformities or soft-tissue deficiencies caused by aging, acne, trauma, surgery, or other causes were included. Persons presenting with connective tissue disorders, skin disorders affecting the face, uncontrolled diabetes mellitus, compromised immune functions or acute inflammatory diseases, known substance abuse, or mental disorders were excluded from participation in the study, as were those who had experienced treatment with any nonabsorbable filler within 6 months before the start of the study. To assess the safety and efficacy of the treatment, follow-up visits were scheduled for day 7, day 28, and 3 months, 6 months, and 12 months after the first injection. Standardized photographs were taken before treatment and during month-3 and month-12 visits. Blood and urine samples were collected and analyzed before injection and at 6 and 12 months after injection. The most common sites of injection were the nasolabial folds (48 percent) and lips (25 percent) followed by glabella folds (8 percent) and other sites. The amount of injected gel ranged from 0.2 to 12 ml. RESULTS: Two hundred twenty-eight patients were followed for 12 months after treatment. In 93 percent, results were judged to be good or very good by the investigators and satisfactory or very satisfactory by the patients. Adverse events that could be ascribed to the treatment were observed in 37 cases, most commonly presenting as transient swelling, hematoma, redness, and pain or itching. In one case, a slight change of skin color at the site of injection was reported. No severe adverse events related to the gel were observed. In one case, neutropenia was detected in month-12 blood samples. The other values were within the normal ranges. CONCLUSIONS: Aquamid is an easy-to-use soft-tissue filler. The effect of tissue enhancement lasted over the entire study period. Transient local tissue reactions observed were attributable to the injection procedure rather than the chemical properties of the gel. During the first year after treatment, no permanent soft-tissue reaction was observed. Long-term results need to be evaluated in the future.

Acrylic Resins↗

PubMed2005

Oxygen consumption in undifferentiated versus differentiated adipogenic mesenchymal precursor cells.

To date, no adequate implant material for the correction of soft tissue defects such as after extensive deep burns, tumor resections or in congenital defects is available. A biohybrid composed of viable adipose precursor cells and an optimised matrix could help towards a solution. Morphologically, preadipocytes resemble fibroblasts and have not yet built a large cytoplasmic lipid droplet as found in differentiated adipocytes. Additionally, preadipocytes are smaller than mature adipocytes allowing a quicker revascularization after transplantation. Furthermore, transplanted preadipocytes can form adipose tissue in vivo whereas the transplantation of mature adipocytes often gives poor results, i.e. oil cysts or shrinkage of the transplant. Since these observations point to differences in metabolic activity between preadipocytes and adipocytes, we investigated the oxygen consumption of preadipocytes stimulated to undergo differentiation, and fibroblasts, by measuring the respiration with a Clark-type oxygen electrode. Preadipocytes had a significantly lower oxygen consumption than mature adipocytes. This advantage in respiration and the better revascularization of undifferentiated adipose tissue cells allow the development of innovative transplants and point to preadipocytes as promising tool to improve transplantations in adipose tissue reconstruction.

3T3 Cells↗

PubMed2005

Implantation of preadipocyte-loaded hyaluronic acid-based scaffolds into nude mice to evaluate potential for soft tissue engineering.

The reconstruction of soft tissue defects following extensive deep burns or tumor resections remains an unresolved problem in plastic and reconstructive surgery since adequate implant materials are still not available. Preadipocytes, immature precursor cells found between mature adipocytes in adipose tissue, are a potential material for soft tissue engineering since they can proliferate and differentiate into adipose tissue after transplantation. In previous studies, we identified hyaluronan benzyl ester (HYAFF 11) sponges to be promising carrier matrices. This study now evaluates, in vitro and in vivo, a new sponge architecture with pores of 400 microm either made of plain HYAFF 11 or HYAFF 11 coated with the extracellular matrix glycosaminoglycan hyaluronic acid. Human preadipocytes were isolated, seeded onto carriers and implanted into nude athymic mice. Explants harvested after 3, 8, and 12 weeks were examined for macroscopical appearance, thickness, weight, pore structure, histology, and immunohistochemistry. Compared to previous studies, we found better penetration of cells into both types of scaffolds, with more extensive formation of new vessels throughout the construct but with only minor adipose tissue. Our encouraging results contribute towards a better seeded and vascularised scaffold but also show that the enhancement of adipogenic conversion of preadipocytes remains a major task for further in vivo experiments.

Adipocytes↗

PubMed2005

Cryopreservation of mature human adipocytes: in vitro measurement of viability.

Cryopreservation of fat grafts for autologous fat transplantation renders repeated harvesting procedures unnecessary. Anecdotic reports have been published, yet data about adipocyte survival are sparse. The beneficial effect of added cryoprotective agents (CPA) is known from other tissues but has not been investigated in adipocytes. Fat cells were harvested using the Coleman method (n = 24). Tests were done after 0, 2, 7, 14, and 30 days of cooling to -20 degrees C and -80 degrees C and after addition of various CPA. Analysis included cell stains, measurement of metabolic activity by MTT and XTT tests, and measurement of cell stability by assessment of extracellular glycerol-3-phosphate-dehydrogenase enzyme. After freezing, up to 92.7% of metabolic activity was lost, but the addition of CPA led to preservation of up to 54% of baseline activity. Also, lower storage temperature showed more cell destruction but yielded higher viability of the surviving cells. Our results implicate that the widely used practice of simple storage in a freezer leads to reinjection of nonviable tissue. Cell survival can be improved by addition of CPA.

Adipocytes↗

PubMed2005

Optimization of the differentiation of human preadipocytes in vitro.

This study aimed at developing an optimal protocol for proliferation and differentiation of preadipocytes that is a prerequisite for constructing an ideal biohybrid composed of viable adipose precursor cells in a three-dimensional matrix. Such an implant could represent an adequate solution for correcting soft tissue defects, e.g., extensive deep burns or tumor resections. Preadipocytes were isolated from human subcutaneous adipose tissue samples and cultured in Dulbecco's modified eagle medium (DMEM)/Ham's F12 medium (F12) or OPTIMEM medium with or without the addition of human serum (hS) or fetal calf serum (FCS). The advantages of fibronectin-coated culture dishes for preadipocyte yield after isolation and differentiation were evaluated. After culture expansion, differentiation was induced by insulin, isobutylmethylxanthine, pioglitazone, dexamethasone, and transferrin in the absence of serum. The extent of differentiation was assayed by measuring the activity of glycerophosphate dehydrogenase as well as counting of differentiated versus undifferentiated cells. Our results show that fibronectin coating does not only strongly increase the yield of preadipocytes after isolation from adipose tissue but also significantly enhances differentiation of precursor cells to mature adipocytes. For optimal cell expansion, DMEM/F12 is more promoting than OPTIMEM and culturing with FCS shows a slightly better proliferation compared with hS supplementation. Differentiation, in contrast, is significantly improved when hS is used instead of FCS during proliferation. Our results smooth the way for autologous preadipocyte culturing and show that hS for preadipocyte culturing opens new and promising perspectives for adipose tissue engineering by optimizing in vitro expansion in cell culture and inducing substantial differentiation.

Adipocytes↗

PubMed2004

Comparison of viable cell yield from excised versus aspirated adipose tissue.

The correction of soft-tissue defects by adipose tissue transplantation often produces poor and unpredictable results. The implantation of isolated and cultured preadipocytes offers a solution to this problem since these cells differentiate into adipocytes when implanted in vivo. A field of major interest is to maximize the yield of preadipocytes isolated from adipose tissue showing only low contamination with other cell types. Aspiration and excision are two concurrent clinical ways of harvesting adipose tissue for the isolation of preadipocytes. This tissue is usually discarded after surgery. In this study, the yield of preadipocytes obtained from liposuction material was compared to that of excised adipose tissue. Furthermore, we determined the loss of precursor cells if isolation of preadipocytes was delayed for 24 h. Preadipocytes were isolated from the stromal cell fraction of human subcutaneous adipose tissue samples. Harvesting of adipose tissue by suction was performed according to the Coleman procedure (manually applied negative pressure using a 10-ml syringe with a blunt tip cannula). Isolation was either carried out within 60 min after extraction or after storage for 24 h in culture medium at 4 degrees C. Isolated preadipocytes were cultured for 24 h, trypsinized and counted in a Neubauer chamber. Our results show clearly that the yield of preadipocytes isolated from liposuction material (within 60 min after extraction and after 24 h of storage) is higher than the cell yield from excised adipose tissue. Overnight storage for 24 h leads to a significant loss of preadipocytes in excised tissue but not in liposuction material. The high yield of cells isolated from liposuction material proves that extraction by suction does not damage the stromal cell fraction in the adipose tissue. If cell isolation is not performed immediately after the operation, liposuction material is clearly the better alternative for storage.

Adipocytes↗

PubMed2003

TIMP-1, MMP-2, MMP-9, and PIIINP as serum markers for skin fibrosis in patients following severe burn trauma.

The wound-healing process of patients with severe burns often leads to the formation of extensive fibrotic scars. In this study, serum concentrations of tissue inhibitors of metalloproteinase-1 (TIMP-1), matrix metalloproteinase-2 (MMP-2), matrix metalloproteinase-9 (MMP-9), and amino-terminal propeptide of procollagen type III (PIIINP) were measured by enzyme-linked immunosorbent assay as markers for excessive cicatrization in 22 patients with acute burn injuries. All patients were followed up for 6 months to determine a fibrotic reaction during the wound-healing process after operative treatment using the Burn Scar Index. Blood samples were drawn immediately before the operation; at postoperative days 1, 3, 7, and 14; and 1, 3, and 6 months after the operation. Twenty patients who underwent elective plastic surgical operations served as the control group. There was a significant increase (p < 0.05) of TIMP-1 in the burned patients by the third postoperative day. Later in the follow-up period, the serum concentrations remained at a significantly elevated level (p < 0.05) compared with preoperative values. In comparison with the control group, the postoperative serum concentrations of TIMP-1 of the burned patients were significantly higher (p < 0.05) at any time and correlated with the total body surface area burned at the third and seventh postoperative days (p < 0.05; r2 = 0.46 versus r2 = 0.53) and the Burn Scar Index after 6 months (p < 0.05; r2 = 0.65). Serum levels of MMP-2 and MMP-9 showed a significant elevation (p < 0.05) only between postoperative days 3 and 14 in patients with burn wounds. PIIINP increased significantly (p < 0.05) in the sera of the burned patients at postoperative day 3 and remained significantly elevated up to 6 months after injury. At any time after trauma, PIIINP serum levels were significantly higher (p < 0.05) in the burned patients than in the control group and correlated with the total body surface area burned at postoperative days 3 and 7 (p < 0.05; r2 = 0.41 versus r2 = 0.44) and the Burn Scar Index after 6 months (p < 0.05; r2 = 0.5). Obviously, the physiological balance between matrix metalloproteinases and their endogenous inhibitors is disturbed after burn trauma. The elevated systemic TIMP-1 concentration might contribute to tissue fibrosis, leading to pathological scar formation. The increase of PIIINP after thermal trauma indicates a fibrogenic component of wound healing.

Biomarkers↗

PubMed2003

Tissue engineering of white adipose tissue using hyaluronic acid-based scaffolds. I: in vitro differentiation of human adipocyte precursor cells on scaffolds.

BACKGROUND AND AIM OF THE STUDY: Reconstruction of soft tissue defects is a challenge in plastic surgery and there is clinical need for adequate solutions. Aim of this study was to develop a biohybrid construct consisting of hyaluronic acid-based scaffolds and human adipocyte precursor cells as a soft tissue filler. METHODS: Human adipocyte precursor cells were obtained by collagenase digestion of adipose tissue samples and seeded on hyaluronic acid-based spongy scaffolds of various degrees of esterification and pore size using different techniques. After cell attachment, adipose differentiation was induced by defined adipogenic factors under serum-free culture conditions. RESULTS: Among the five different scaffold types under investigation the highest cell attachment rate was observed for the HYAFF scaffold with 100% esterification and a mean pore size of 400microm (HYAFF 11lp). For inoculation of human adipocyte precursor cells on hyaluronic acid-based scaffolds a "drop-on" technique and low-pressure centrifugation using a Speed Vac airfuge were compared. With respect to efficacy, cell distribution and simpleness the drop-on method proved to be the method of choice. In a serum-free medium supplemented with 66nM insulin, 100nM cortisol and 1microg/ml troglitazone a substantial proportion of cells underwent adipose differentiation as assessed by lipid accumulation and emergence of glycerol-3-phosphate dehydrogenase activity, a lipogenic marker enzyme. CONCLUSION: Hyaluronic acid-based scaffolds appear to be a suitable three-dimensional carrier for the culture and in vitro differentiation of human adipocyte precursor cells.

Adipocytes↗

PubMed2003

Pathogenic role of interleukin-6 in the development of sepsis. Part I: Study in a standardized contact burn murine model.

OBJECTIVE: To establish a representative model for the evaluation of interleukin (IL)-6 and IL-6 receptor for pathogenicity and lethality in the postburn period. DESIGN: Ten-week-old C 57 BL/6J mice received a 20% body surface area contact burn and/or lipopolysaccharide (LPS) 48 hrs later. Standardized burns were created with a metal stamp of 150 degrees C of defined pressure and surface area (2.4525 Newton/0.00166 m2) over a period of 11 secs. The depth of dermal injury was verified histologically. The following groups were formed: I: no burn, no LPS (n = 35); II: burn, no LPS (n = 140); III, no burn, LPS (n = 56); and IV, burn, LPS (n = 80), to study the effect of burn alone, sepsis alone, or the combination. Lethal LPS dose (LD100) was determined by application of LPS in increasing doses (200, 300, 400, and 500 microg, n = 32) after burns. MEASUREMENTS: Concentrations of interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma), and leukocytes, platelets and organ pathology were evaluated. SETTING: Research laboratory. RESULTS: Burn and LPS showed an additive effect on the release of IL-6 but not of TNF-alpha and IFN-gamma. Leukocyte and platelet numbers decreased significantly (group IV) compared with the other groups (I-III). The maximal levels of IL-6 in group IV were reached earlier than those of TNF-alpha. The contact burn model has a mortality rate of 30%, which is close to clinical outcome. We found the model of contact burn superior to scald or flame burn models. A dose of 400-microg LPS was found to be the lethal LPS dose (LD100). CONCLUSIONS: Our data suggest that preexisting burn injury increases the response to endotoxin. TNF-alpha is not involved in priming. IL-6 on the other hand is a very representative parameter for priming. Because TNF-alpha was obviously not the causative factor, it was concluded that the application of anti-IL-6-mAb should be of great value. Therefore, a therapeutic application was designed, see part II.

Animals↗

PubMed2003

Pathogenic role of interleukin-6 in the development of sepsis. Part II: Significance of anti-interleukin-6 and anti-soluble interleukin-6 receptor-alpha antibodies in a standardized murine contact burn model.

OBJECTIVE: The in vivo effects of anti-interleukin-6 (anti-IL-6) and anti-interleukin-6-alpha receptor (anti-IL-6R) monoclonal antibodies on immune response and survival rate of a burn with subsequent infection were assessed. SUBJECTS: Ten-week-old C 57 BL/6J mice received a standardized contact burn; 48 hrs later endotoxin (LPS) was injected intraperitoneally to induce systemic inflammation. Ten different groups were studied. Groups I-IV sustained a burn and/or a LPS-stimulus but did not receive any anti-cytokines and served as controls. Treatment groups V-X sustained the same injuries but also received anti-IL-6 and anti-IL-6R intravenously either before or after the LPS stimulus. In a further part of the study, a lethal dose of LPS was injected (LPS-LD(100) group) followed by an injection of anti-IL-6 antibody and/or anti-IL-6R antibody. MEASUREMENTS: Serum concentrations of IL-6, tumor necrosis factor (TNF)-alpha, interferon (IFN)-gamma, and white blood cell and platelet counts were determined, and the survival rate over a 2-wk period was assessed. RESULTS: Treatment with anti-IL-6 slightly decreased the inflammatory response when it was given before or after LPS application. The inflammatory response was not decreased after treatment with anti-IL-6R. In the groups that received a combination of anti-IL-6 and anti-IL-6R, there was a significant reduction of the inflammatory response. This was more pronounced when the anti-cytokines were applied after LPS application. A significant reduction in mortality could be shown with both antibodies in the treatment groups and the groups that had received a lethal dose of LPS (LPS-LD(100) group). CONCLUSIONS: IL-6 has a low inflammatory potential, and IL-6R has no inflammatory potential by itself. In contrast, the IL-6/IL-6R complexes have a higher inflammatory potential. Mortality could be reduced by each antibody alone as well as by the combination, supporting the hypothesis that the inflammatory and lethal potentials of IL-6 are not identical. The study suggests that the use of antibodies against IL-6 or IL-6R is effective in the prevention of systemic inflammation in a murine burn model.

Animals↗

PubMed2003

Ball lightning burn.

Ball lightning is a rare physical phenomenon, which is not yet completely explained. It is similar to lightning but with different, peculiar characteristics. It can be considered a mix of fire and electricity, concentrated in a fireball with a diameter of 20-cm that most commonly appears suddenly, even in indoor conditions, during a thunderstorm. It moves quickly for several meters, can change direction, and ultimately disappears. During a great storm, a 28-year-old man and his 5-year-old daughter sustained burn wounds after ball lightning came from the outdoors through a chimney. These two patients demonstrated signs of fire and electrical injuries. The father, who lost consciousness, sustained superficial second-degree burn wounds bilaterally on the zygomatic area and deep second-degree burn wounds on his right hand (total body surface area, 4%). His daughter demonstrated superficial second-degree burn wounds on the left part of the face and deep second-degree and third-degree burn wounds (total body surface area, 30%) on the left neck, both upper arms, and the back. In this article, the authors report the first two cases of burn injuries resulting from ball lightning contact indoors. The literature on this rare phenomenon is reviewed to elucidate the nature of ball lightning. Emphasis is placed on the nature of injuries after ball lightning contact, the therapy used, and the long-term complications.

Adult↗