Ideal Time Point Research Articles

Intraperitoneal Administration of Bevacizumab Intraoperatively Does Not Affect Abdominal Wound Healing in Rats

Background: Bevacizumab is a monoclonal antibody targeted at vascular endothelial growth factor (VEGF) to treat advanced colorectal cancer as well as other malignancies, but the ideal time point for its administration in patients scheduled for surgery is not well defined due to serious concerns regarding possible side effects on wound healing. Therefore, we conducted an experimental study in rats to clarify this issue. Methods: Four groups of 10 Wistar rats each underwent a 4-cm midline laparotomy and closure of the wound in 2 layers. In the treatment groups (A and B), bevacizumab (Avastin®) received a single dose of 5 mg/kg i.p., and an equal amount of saline was given to the control groups (C and D). Groups A and C were sacrificed on the 7th postoperative day, and groups B and D on the 14th postoperative day. Wounds were inspected by two independent observers upon sacrifice and results were recorded; wound tissues were sent for histology to assess the degree of fibrosis and measurement of tissue hydroxyproline levels. Serum levels of endothelin-1, C-reactive protein, pro-oxidant/antioxidant balance and carbonylated proteins were also determined. For statistical analysis, the Mann-Whitney U test was used. Results: Wound healing did not differ among groups both on the 7th and the 14th postoperative days, and there was also no significant difference regarding the degree of inflammation, fibroblast proliferation and collagen synthesis, as well as hydroxyproline and biochemical marker levels among the groups. Conclusions: Intraperitoneal bevacizumab administered intraoperatively does not significantly affect abdominal wound healing in rats.

Gemcitabine depletes regulatory T‐cells in human and mice and enhances triggering of vaccine‐specific cytotoxic T‐cells

Particle-mediated epidermal delivery (PMED) is a potent genetic vaccination method. However, a recent report found PMED only poorly and infrequently triggered antigen-specific cytotoxic T-cells in cancer patients. Here, we show that injection of the chemotherapeutic drug Gemcitabine in mice results in improvement of the efficacy of subsequent PMED vaccination against NY-ESO-1. We found in mice and in cancer patients that administration of Gemcitabine induces a transient reduction in the percentage of regulatory T-cells among CD4-positive cells. The higher relative sensitivity of regulatory T-cells compared to other CD4-positive T-cells toward cytostatic drugs can be linked to the higher frequency of proliferating cells in the regulatory compartment compared to the nonregulatory CD4-compartment in healthy people and cancer patients. Thus, by affecting regulatory T-cells more than other lymphocyte subsets, chemotherapeutic agents can create a transient hyperimmunoreactive window. Such a window would provide an ideal timepoint to administer a vaccine expected to induce a therapeutically relevant anticancer cytotoxic T-cell response.

Diagnostics, treatment, and follow-up in craniopharyngioma.

Craniopharyngiomas are partly cystic embryogenic malformations of the sellar and parasellar region, with up to half the 0.5–2.0 new cases per million population per year occur in children and adolescents. Diagnosis profile for pediatric and adult craniopharyngioma is characterized by a combination of headache, visual impairment, and polyuria/polydipsia, which can also include significant weight gain. In children, growth retardation, and/or premature puberty often occur later or postoperatively. Recommended therapy with favorable tumor localization is complete resection; with unfavorable tumor localization (optic nerve and/or hypothalamic involvement), consensus is still pending whether a limited resection followed by local irradiation is more prudent. Even though overall survival rates are high (92%), recurrences after complete resection and progressions after incomplete resection can be expected. Accordingly, a randomized multinational trial (KRANIOPHARYNGEOM 2007) has been established to identify optimal diagnosis, treatment (particularly the ideal time point of irradiation after incomplete resection), and quality of life strategies of this chronic disease – most notably the morbid hypothalamic obesity in ∼50% of long-term survivors. We report on craniopharyngioma origins, its pathological manifestations, and specific challenges these sequelae pose regarding diagnosis, treatment, and life-long multi-discipline quality of life management for both adult and childhood craniopharyngioma patients.

Dynamic change of mother-source neutralizing antibodies against enterovirus 71 and coxsackievirus A16 in infants

Background Enterovirus 71 (EV71) and coxsackievirus A16 (Cox A16) are major causative agents for hand, foot and mouth disease (HFMD). Studies indicate that the frequent HFMD outbreaks result in a few hundreds children's death in China in recent years. The vaccine and other research for HFMD need to be developed urgently. The aims of our study were: to explore dynamic development of mother-source neutralizing antibodies against EV71 and Cox A16 in infants from Jiangsu Province, China, and to provide the fundamental data for further establishing of corresponding immunization course. Methods Peripheral blood samples were collected from 133 of parturient women once immediately before delivery and their infants at two and seven months of age. Method of micro-dose cytopathogenic effect was used to measure neutralizing antibodies against EV71 and Cox A16, respectively. Results Seropositive rates of anti-EV71 and anti-Cox A16 in prenatal women were 79.7% (106/133) and 92.5% (123/133), respectively; geometric mean titers (GMTs) were 29.0 and 61.9; 75.9% (101/133) prenatal women were both positive in anti-EV71 and anti-Cox A16; seropositive rates of anti-EV71 and anti-Cox A16 were 25.6% (34/133) and 38.3% (51/133) in infants at two months of age; GMTs were 12.3 and 18.0, respectively. GMTs of anti-EV71 were significantly higher for infants at seven months (82.6) compared with that at two months (P <0.05), showing infants had inapparently infected by EV71 during two to seven months. Although only one offspring (0.75%) at seven months was found having anti-Cox A16 transfered from maternal, this observation suggested no maternal antibody may remain in infants at seven months. Conclusions The prevalence of EV71 and Cox A16 were relatively high in Jiangsu Province. Bivalent vaccine against both EV71 and Cox A16 should be developed, and the ideal time point for prime immunization for infants is around 2–5 months of age.

Management of Prenatally Diagnosed Abdominal Lymphatic Malformations

Abdominal lymphatic malformations (ALM) are rare congenital malformations that can regress spontaneously or lead to serious complications. Thus, the appropriate management may be challenging, particularly since pertinent literature is missing. We present our experience in the management of 5 patients with prenatally diagnosed ALM and their outcome and propose a decision-making algorithm. We retrospectively reviewed the history, diagnostics, therapy, complications, and outcome of 5 patients with a prenatal diagnosis of ALM, referred to our department between January 2006 and February 2008. ALM was prenatally diagnosed by ultrasound in all patients (gestational age 21, 23, 23, 32, and 34 weeks). MRI was performed pre- and postnatally in one patient and postnatally in another. Clinical symptoms ranged from none to respiratory distress and abdominal compartment syndrome. One ALM involuted. 2 patients underwent primary OK-432 treatment. This led to a 70% size reduction in one patient. The other developed massive intracystic bleeding and required emergency surgery. 2/3 patients with surgery needed segmental bowel resection and 3/3 stayed recurrence-free. Complications included one partial inferior vena cava thrombosis after surgery, one subileus, and one hemorrhage after OK-432 application. Asymptomatic and regressing ALM are best managed conservatively ("watchful waiting") while symptomatic ALMs require surgery. Further studies are necessary to determine the ideal timepoint for intervention for non-regressing ALM.

Which is the ideal point of time to perform intraoperative 3D imaging in dorsal stabilisation of thoracolumbar spine fractures? A matched pair analysis

Introduction After dorsal stabilisation of vertebral fractures by an internal fixateur the postoperative computed tomography is a standard procedure to control the positions of the pedicle screws, the success of the reposition, the clearance of the spinal canal and to plane an additive secondary ventral stabilisation. An intraoperative scan with a 3D image intensifier may clarify these questions directly after the implantation with the possibility of an immediate correction of the implants. The aim of this study was to find out the optimal point of time to perform an intraoperative 3D scan and if a postoperative computed tomography is dispensable. Patients and methods Intraoperative 3D scans were carried out on 33 patients with thoracolumbar spine fractures (T11-L5) after bi-segmental fixateur interne montage (Group 1). A matched pair group of 33 patients (Group 2) with a 3D scan after implantation of pedicle screws was built. A postoperative computed tomography of the instrumented spinal section was done in all patients. The following measurements were done in sagittal and axial reconstruction planes and were compared: classification of screw positions, maximal axial diameter of pedicles, cortical perforation of the screws. Additionally in Group 1 the distance between the upper and lower end plates of the injured section, the height of posterior vertebral body wall, the dislocation of the posterior wall and the minimal diameter of the spinal canal were measured. Results The intraoperative scoring of pedicle screws positions and the measurement of pedicle width showed in both groups a significant accordance with the computed tomography determinations. The measurements “posterior wall dislocation” and “diameter of spinal canal” were only possible in 24 3D scans and showed a significant difference compared with the CT data. The picture quality in Group 2 was scored significantly better than for Group 1 with the complete assembly of the fixateur. Conclusion The ideal point of time for an intraoperative 3D imaging with the present intensifier generation is directly after pedicle screw insertion. The reliable determination of the spinal canal diameter, of posterior wall fragments and of the exact fracture morphology is only possible by postoperative computed tomography.

63 EXPERIMENTALLY INDUCED EARLY DEGENERATION OF THE HIP JOINT IN A SHEEP MODEL

Purpose: A non-spherical femoral head is a known risk factor for early osteoarthritis of the hip joint. Recently, a new pathomechanism termed femoro-acetabular impingement has been postulated as an underlying cause. This mechanism is postulated to represent an abutment conflict between the non-spherical portion of the head/neck junction and the acetabular rim during extreme flexion and internal rotation. It is thought to lead to repetitive trauma resulting in subsequent lesions of the labrum and adjacent cartilage at the acetabular rim or femoral head. The aim of this research study is to confirm in an animal model, that a non-spherical head contour leads to primary lesions at the acetabulum and at the nonspherical portion of the femoral head similar to the findings in human hip joints. Methods: Sheep have been chosen as animal model since the acetabular anatomy is close to that in humans including the presence of a labrum and the femoral head has a distinct non-spherical extension of the cartilage in its superior portion. By performing an intertrochanteric adduction osteotomy of the proximal femur the non-spherical portion was rotated toward the weight bearing zone and created similar impingement conditions during the end range of flexion as postulated in young adult human beings. In a pilot study with 6 sheep localized peripheral hip joint lesions could be produced at the acetabular rim and the non-spherical portion of the head similar to the finding in humans. For this study a total of 16 sheep was chosen. The sheep were divided into 4 groups, each group with 4 sheep that were sacrificed after 14, 22, 30, 38-weeks after surgery. After sacrifice the joint cartilage was evaluated for visible alterations macroscopically according to grading by Beck et al. Thereafter samples were harvested of standardized segments on both, the operated and non-operated (control) sides for histological work-up using conventional staining methods with Toluidin-blue O to evaluate proteoglycan depletion and grading of cartilage alterations according to Mankin. Results: Localized labral lesions as also typically seen in humans were present in 14 out of 16 hips on the operated side versus 2 lesions on the unoperated side. Cartilage lesions including flaps, tears or superficial fraying were seen in all 16 operated hips in the caudodorsal segment of the acetabulum versus alterations in 3 out of 16 hips on the unoperated side (different stages of alteration showed in Figure). Histochemical staining confirmed the macroscopic findings and showed statistically significant higher degenerative changes according to the Mankin staging in the caudodorsal segment on the operated side versus the non-operated side. Conclusions: With the presented animal model a cam impingementmechanism and localized, peripheral intraarticular lesions of the hip similar those seen in humans could be reproduced. Therefore a model of development of early osteoarthritis in the hip with different stages of the disease could be defined. This allows to develop and to evaluate new invasive and non-invasive treatment methods for this pathomechanism. It will be also used to further clarify the sequential steps of the osteoarthritic process and help to define the ideal time point for surgical interventions.

Optimierung des perioperativen Managements am Beispiel der laparoskopischen Leistenbruchoperation

Although about 150,000-200,000 hernia repair procedures are performed every year in Germany alone, fast-track concepts are mainly ignored for this type of surgery. Therefore, this study was designed to analyze the perioperative management of patients undergoing laparoscopic hernia repair, performed as transabdominal preperitoneal patch hernioplasty (TAPP). Based on these results, the clinical management was optimized in order to shorten the length of stay without affecting the quality of treatment or the complication rate, and the effects of the optimization strategies were reanalyzed. With ethics committee approval 249 patients undergoing TAPP procedures were investigated. In the first two study sections, problems of the perioperative management were identified first retrospectively (n=129) and then prospectively (n=60). Based on these results, the clinical management was then redesigned and optimized. A TAPP score was developed including the parameters age, ASA physical status, extent and severity of the procedure and postoperative pain level. Patients were scored 24 h after surgery, and clear-cut criteria were defined for discharge home on the second postoperative day. Furthermore, all patients received 10 mg of sodium picosulfate to avoid postoperative constipation. In the third study section (n=60) the impact of the optimization strategies on length of stay, need for pain medication, complication rate and patient satisfaction (based on the PPP33 questionnaire) was evaluated. Patients were on average approximately 60 years old (range 22-92 years), and demographic data were comparable within the study sections. As a result of the optimization process, 72% of the patients could be discharged home on the second postoperative day whereas previously that had only been possible in 5%. Accordingly, the postoperative length of stay (including the day of surgery) was significantly reduced from 4.2+/-0.6 to 3.3+/-0.6 days. By the administration of sodium picosulfate, 92% of all patients reported defecation on the first day after surgery, whereas previously the majority of patients (60%) had complained of constipation at this time. No serious complications were observed. The number of minor complications was very low in total and neither complication rate nor patient satisfaction was affected by earlier discharge. The second day after surgery was judged to be the ideal time point for discharge by 81% of all patients. This study demonstrates that the length of stay after laparoscopic hernia repair can be reduced by one day by relatively simple means without affecting patient satisfaction or the complication rate. Most important is the introduction of clear-cut criteria that allow an early decision-making for discharge home. Moreover, many patients complain of constipation after laparoscopic surgery, and this may prolong the length of stay. This problem can be solved completely by the standardized use of sodium picosulfate, administered on the evening after surgery.

Telemetric monitoring of tracheal pressure after tracheal occlusion for treatment of severe congenital diaphragmatic hernia

Prenatal tracheal occlusion using endoscopic techniques obstructs the normal egress of lung fluid during pulmonary development and stimulates lung growth in cases of congenital diaphragmatic hernia (CDH). Although FETO might be an effective strategy for treatment of CDH, the mechanism especially due to the supposed increasing transpulmonary pressure is unknown. The purpose of this study was to monitor the pressure below the attached balloon in the fetal lamb telemetrically. Four time-dated pregnant Merino ewes underwent fetal and maternal surgery. A special prepared silicone catheter was placed below the epiglottis by laryngoscopy on day 110 or 140 of gestation. The tracheal pressure below the fixed catheter could be monitored telemetrically using the Data Sciences TA11-PA-C40 pressure device. Hundred and twenty measurement points were recorded over a period of 2 min. A maximum of lung pressure rate was found immediately after implantation (23.7 +/- 4.6 mm Hg). During the first hour, the pressure decreased to an average value of 16.9 mmHg. About 70 h after the block, this value decreased to a minimum level of 8.3 +/- 0.4 mmHg. Decreasing pressure variation might indicate that lung growth has stopped and that the ideal point of time to remove the balloon is achieved. Increasing pressure has to be related to the morphometric analysis of the lung's structural development and maturation, comparing the efficacy of FETO in preventing or reversing pulmonary hypoplasia. Further investigation of continuous telemetric monitoring of tracheal pressure in the fetal lamb is required.

Successful doxycycline treatment of lymphatic fistulas: report of five cases and review of the literature

Lymphatic fistulas are commonly known complications after lymph node or soft tissue resection, requiring long and sometimes difficult therapy. Doxycycline is an antibiotic agent, which was used in the treatment of pneumothoraces and pleural effusions. The aim of the study was to evaluate the effect of local doxycycline instillation on postoperative lymphatic effusions. Five patients with postoperative lymphatic fistulas of different locations (inguinal, gluteal, and thoracic) were included. Patients received local doxycycline instillation after a postoperative observation period of 19-27 days without spontaneous closure of the fistulas. Postoperative lymphatic fistulas closed after a single injection of doxycycline in four out of five patients within 2 days, in one patient, injection had to be repeated to achieve closure of the fistula. Drain removal was possible within 4 days after instillation in all patients; no complications occurred. Doxycycline can be used for treatment of persistent postoperative lymphatic fistulas in different locations. In further studies, this therapeutic concept should be evaluated with larger numbers of patients with regard to the ideal point of time, frequency, and mode of application.

Characterization of the Secreted Proteome of Rat Hepatocytes Cultured in Collagen Sandwiches

Analysis of proteins in biological samples opens up the possibility of discovering new markers of toxicity. The liver is one of the primary targets of drug-induced toxicity, and it also secretes many plasma proteins, which can be measured clinically. Most of the plasma proteins produced by the liver are secreted by hepatocytes, but there is little information regarding the protein profile secreted by these cells. The purpose of this study was to analyze the secreted proteome of primary rat hepatocytes in a collagen gel sandwich configuration by a gel-LC-MS/MS procedure. We identified over 600 peptides corresponding to more than 200 proteins. The protein profile included over 50 plasma proteins, suggesting that the cultured hepatocytes secrete many of the proteins that they produce in vivo. Our data also suggests that the hepatocytes are actively remodeling their environment, since we identified several structural extracellular matrix proteins as well as some proteins known to be secreted specifically during liver regeneration. We also identified two proteins, alpha1-antitrypsin and alpha2-macroglobulin, whose secretions appear to be down-regulated in cells exposed to aflatoxin B1. It was noted that a 15 nM dose of aflatoxin B1 led to substantially diminished levels of these proteins and that day 6 of incubation was the ideal time point for medium collection. These data suggest that proteins in the conditioned medium of hepatocyte sandwich culture might lead to the discovery of biomarkers for drug-induced chemical toxicity.

A systematic evaluation of bioelectrical impedance measurement after hemodialysis session

There is still no definitive indication about the ideal point of time to perform bioimpedance analysis (BIA) in hemodialysis patients. Furthermore, the interpretation of data in this regard is difficult because there is still no comprehensive information about the fluctuations in BIA variables occurring in these subjects. The aim of this study was to assess BIA changes occurring in hemodialysis and specifically in the dry-weight state. We studied 27 anuric patients (20 males and 7 females; age 56.1 +/- 13.7 years) on chronic hemodialysis. Single-frequency BIA (R, resistance; Xc, reactance; and PhA, phase angle) was performed (1) before and at the end of hemodialysis (dialysis period); (2) 15, 30, 60, 90, and 120 minutes after hemodialysis (postdialysis period); and (3) 24, 48, and 68 hours after hemodialysis (interdialysis period). Body weight decreased by 2.8 +/- 0.8 kg during hemodialysis, was unchanged during the postdialysis period, and progressively rose during the interdialysis period. At the same time, BIA variables significantly increased during hemodialysis (R, 453 +/- 74 and 542 +/- 98 ohm; Xc, 38 +/- 10 and 53 +/- 16 ohm; P < 0.05), remained stable over the 120-minute period after treatment (R, 538 +/- 94, 539 +/- 95, 538 +/- 94, 541 +/- 95, and 544 +/- 95 ohm; and Xc, 53 +/- 15, 53 +/- 15, 51 +/- 16, 52 +/- 16, and 52 +/- 16 ohm; NS), and subsequently declined [R, 471 +/- 79 (P= <0.05 vs. postdialysis), 449 +/- 71 (P= <0.05 vs. postdialysis), 424 +/- 68 (P= <0.05 vs. postdialysis) ohm; Xc, 42 +/- 13 (P= <0.05 vs. postdialysis), 37 +/- 10 (P= <0.05 vs. postdialysis), 34 +/- 13 (P= <0.05 vs. postdialysis) ohm]. The stability of BIA measures during postdialysis was confirmed by the constant relationship found between R/height and Xc/height. Also PhA increased after dialysis (4.8 +/- 1.1 degrees vs. 5.7 +/- 1.3 degrees, P < 0.05), was unchanged during the following 120 minutes and decreased in the interdialysis period (5.1 +/- 1.3 degrees, 4.8 +/- 1.0 degrees, and 4.5 +/- 1.1 degrees, P < 0.05). At the end of hemodialysis and during the postdialysis period total body water (TBW) estimated from BIA was similar on average to TBW calculated using Watson formulas (37.2 +/- 6.3 L vs. 36.2 +/- 5.7 L, NS). On the contrary, when patients were hyperhydrated BIA significantly overestimated the Watson's values. In hemodialysis patients BIA variables fluctuate to a considerable extent (with the highest values immediately after hemodialysis), but remain constant and highly reproducible over the 120 minutes after the end of hemodialysis, that is, in a dry-weight state. Thus, taking into consideration that the point in time chosen for performing BIA is crucial to properly assess body composition, BIA can be appropriately performed at anytime during the postdialysis period, provided that hydration status does not change due to food or drink consumption.

A Mixture Model to Assess The Effect of Hormonal Stimulation on the Development of Follicles in Prepubertal Heifers

Summary The goal of this study is to assess the effect of follicle stimulating hormone (FSH) on the growth of follicles in prepubertal heifers. Adams and co-workers have shown that follicles develop in waves as in post-pubertal heifers: after a few days, a dominant follicle arises on one of the two ovaries whereas the other subordinate follicles regress. The dominant follicle remains detectable when the next wave occurs. The number and the diameters of these follicles have been measured for 14 days. The same experiment was repeated after a 3-day FSH stimulation. Several large follicles now arise simultaneously on both ovaries together with small follicles which cannot be detected any more when the next wave occurs. Unfortunately, follicles were not individually tracked. Therefore a three-component mixture model was used to describe the potential simultaneous observation of dominant, decaying dominant and subordinate follicles before the FSH stimulation; the same type of model was considered for large, decaying large and small follicles after the hormonal stimulation. Moreover, for technical reasons, only follicles larger than 2 mm are observable. Measurements were thus left truncated. The models fitted are used to quantify the effects of the stimulation and to determine the ideal time point at which follicles should be punctured under ultrasonography to harvest ovocytes that will be fertilized and cultured in vitro.

Post-stroke depression: diagnosis and therapy

Depression occurs at least temporarily in up to 30 - 40 % of all patients that have experienced a stroke. In the literature the term of a "Poststroke Depression" (PSD) has already been established. However standardised criteria for this diagnoses do not exist to this day. In most cases the DSM or ICD classification system is applied. Other investigators used various psychiatric rating scales. Even though it is generally acknowledged that there is a high prevalence, the occurrence of depression in combination with a stroke fails in most cases to be diagnosed or is left untreated. The need for treatment is even more pronounced by studies showing that a combination of stroke and depression will result in a less favourable outcome, particularly pertaining to that of functional treatment (motor skills, independent participation in activities of daily life). This difference in impairment between a depressed stroke patient and a not depressed stroke patient could be proven in studies that have been conducted over years. Despite the necessity of treatment that can be concluded from such a finding, the recent literature does not offer consistent information as to the ideal point of time for intervention nor the kind and intensity necessitated. According to singular studies, that have rarely been conducted under controlled conditions, there have been positive outcomes after early treatment. Along with psychostimulants that were most predominantly applied in the USA, as well as conventional tricyclic antidepressants, the group of the "Selective Serotonin Reuptake Inhibitors, SSRIs" have resulted in particularly favourable clinical outcomes.

Effect of FLT3 Ligand and Granulocyte Colony-Stimulating Factor on Expansion and Mobilization of Facilitating Cells and Hematopoietic Stem Cells in Mice: Kinetics and Repopulating Potential

We have previously identified a cellular population in murine bone marrow that facilitates engraftment of highly purified hematopoietic stem cells (HSC) across major histocompatibility complex (MHC) barriers without causing graft-versus-host disease. Here we investigated the effect of flt3 ligand (FL) and granulocyte colony-stimulating factor (G-CSF) on the mobilization of facilitating cells (FC) and HSC into peripheral blood (PB). Mice were injected with FL alone (day 1 to 10), G-CSF alone (day 4 to 10), or both in combination. The number of FC (CD8+/βTCR−/γδTCR−) and HSC (lineage−/Sca-1+/c-kit+) was assessed daily by flow cytometry. Lethally irradiated allogeneic mice were reconstituted with PB mononuclear cells (PBMC). FL and G-CSF showed a highly significant synergy on the mobilization of FC and HSC. The peak efficiency for mobilization of FC (21-fold increase) and HSC (200-fold increase) was reached on day 10. Our data further suggest that the proliferation of FC and HSC induced by FL in addition to the mobilizing effect mediated by G-CSF might be responsible for the observed synergy of both growth factors. Finally, the engraftment potential of PBMC mobilized with FL and G-CSF or FL alone was superior to PBMC obtained from animals treated with G-CSF alone. Experiments comparing the engraftment potential of day 7 and day 10 mobilized PBMC indicate that day 10, during which both FC and HSC reached their maximum, might be the ideal time point for the collection of both populations. © 1998 by The American Society of Hematology.

Effect of FLT3 Ligand and Granulocyte Colony-Stimulating Factor on Expansion and Mobilization of Facilitating Cells and Hematopoietic Stem Cells in Mice: Kinetics and Repopulating Potential

AbstractWe have previously identified a cellular population in murine bone marrow that facilitates engraftment of highly purified hematopoietic stem cells (HSC) across major histocompatibility complex (MHC) barriers without causing graft-versus-host disease. Here we investigated the effect of flt3 ligand (FL) and granulocyte colony-stimulating factor (G-CSF) on the mobilization of facilitating cells (FC) and HSC into peripheral blood (PB). Mice were injected with FL alone (day 1 to 10), G-CSF alone (day 4 to 10), or both in combination. The number of FC (CD8+/αβTCR−/γδTCR−) and HSC (lineage−/Sca-1+/c-kit+) was assessed daily by flow cytometry. Lethally irradiated allogeneic mice were reconstituted with PB mononuclear cells (PBMC). FL and G-CSF showed a highly significant synergy on the mobilization of FC and HSC. The peak efficiency for mobilization of FC (21-fold increase) and HSC (200-fold increase) was reached on day 10. Our data further suggest that the proliferation of FC and HSC induced by FL in addition to the mobilizing effect mediated by G-CSF might be responsible for the observed synergy of both growth factors. Finally, the engraftment potential of PBMC mobilized with FL and G-CSF or FL alone was superior to PBMC obtained from animals treated with G-CSF alone. Experiments comparing the engraftment potential of day 7 and day 10 mobilized PBMC indicate that day 10, during which both FC and HSC reached their maximum, might be the ideal time point for the collection of both populations. © 1998 by The American Society of Hematology.