Cocultivation of umbilical cord blood cells with endothelial cells leads to extensive amplification of competent CD34+CD38- cells

Objective: In this report, methods to expand the number of human cord blood hematopoietic stem cells were explored. MATERIALS AND METHODS: CD34+ cord blood cells were expanded in the presence of various cytokine combinations in either a stroma-free cell culture system or a preformed porcine microvascular endothelial cell layer. After 7 to 21 days, stem cell number and function were monitored. In addition, the replicative history of stem cells was tracked using the fluorescent dye, PKH26. RESULTS: With the addition of various cytokine combinations, total cellular expansion was equivalent for both culture systems, although the endothelial cell-based system contained statistically greater numbers of CD34+ cells. By day 21, the endothelial-based system receiving the FLT3L, SCF, IL-6, and GM-CSF cytokine combination contained five-fold greater numbers of CD34+ than the stroma cell-free culture cell system. Endothelial-based cultures receiving these four cytokines plus megakaryocyte growth and development factor produced a 640-fold expansion of CD34+CD38- cells as compared to a four-fold expansion in the stroma-free system. The number of progenitor cells generated was similar with both systems, yet the greatest degree of expansion of cobblestone area-forming cells was observed in the endothelial based cultures (11-fold vs four-fold). Virtually all CD34+ and CD34+CD38+ cells expanded in the presence of endothelial cells had undergone self replication by day 10, yet stromal cell-free cultures contained a significant number (4.8%) of quiescent cells. Identical numbers of re-isolated cord blood CD34+ cells expanded in both systems exhibited a similar ability to engraft and generate cells belonging to multiple hematopoietic lineages in human fetal bones implanted in immunodeficient mice. CONCLUSIONS: These results suggest that the use of preformed endothelial cell monolayers might permit the ex vivo generation of sufficient numbers of cord stem cells to serve as successful grafts for adult transplant recipients.     

Comparison of budesonide and beclomethasone dipropionate for treatment of asthma.

Beclomethasone dipropionate (BDP) and budesonide (BUD) were each given in a dose of 200 micrograms twice daily by metered dose inhaler to 10 asthmatic children already dependent on treatment with steroids. In a double blind randomised crossover study each course lasted one month. No clinically important differences were found between the two treatments when symptom scores, symptom free days, additional use of salbutamol, and results of lung function tests were considered. Metyrapone mildly reduced the plasma concentration of 11-deoxycortisol in two patients during treatment with budesonide, and in four during treatment with beclomethasone. It is concluded that although they are usually safe, both drugs may cause mild adrenal suppression when given in a dose of 200 micrograms twice daily.     

Direct correlation between plasma renin activity and severity of the ovarian hyperstimulation syndrome

Plasma renin activity (PRA) and aldosterone were measured throughout the luteal phase in 21 anovulatory patients who developed ovarian hyperstimulation syndrome (OHSS) during menotropin induction of ovulation. The pattern of PRA in hyperstimulated cycles is characterized by a midluteal peak, which declines to normal in the late luteal phase in nonconceptual cycles, whereas a sustained elevation of PRA occurs in conceptual cycles. Midluteal PRA is significantly (P less than 0.001) elevated in patients with OHSS compared with controls. In the mild form of the disease, the median of PRA is 7.5 (range 6 to 11) ng angiotensin I (AI)/ml/hr, significantly higher than the median for controls, 3.0 ng AI/ml/hr (range, 1.4 to 5). In moderate OHSS, PRA was 24.5 (range, 10 to 40) ng AI/ml/hr, whereas, in the severe form of OHSS, PRA was 55.0 (range, 29 to 95) ng AI/ml/hr. A significant correlation (P less than 0.05) was demonstrated between PRA and either progesterone or 17 beta estradiol (E2). The renin-angiotensin cascade is implicated in new vessel formation. Angiogenesis itself is associated with a rapid increase in capillary permeability. The recent demonstration of high plasma renin-like activity in human follicular fluid and the present observation of high PRA in patients with OHSS may imply that the locally active renin angiotensin system, through induction of new vessel formation and increase in capillary permeability, may have a casual relationship to the ovarian enlargement and extracellular fluid accumulation that are the hallmarks of OHSS.     

Effect of the Total Facet Arthroplasty System after complete laminectomy-facetectomy on the biomechanics of implanted and adjacent segments

Background contextLumbar fusion is traditionally used to restore stability after wide surgical decompression for spinal stenosis. The Total Facet Arthroplasty System (TFAS) is a motion-restoring implant suggested as an alternative to rigid fixation after complete facetectomy.PurposeTo investigate the effect of TFAS on the kinematics of the implanted and adjacent lumbar segments.Study designBiomechanical in vitro study.MethodsNine human lumbar spines (L1 to sacrum) were tested in flexion-extension (+8 to ?6 Nm), lateral bending (±6 Nm), and axial rotation (±5 Nm). Flexion-extension was tested under 400 N follower preload. Specimens were tested intact, after complete L3 laminectomy with L3–L4 facetectomy, after L3–L4 pedicle screw fixation, and after L3–L4 TFAS implantation. Range of motion (ROM) was assessed in all tested directions. Neutral zone and stiffness in flexion and extension were calculated to assess quality of motion.ResultsComplete laminectomy-facetectomy increased L3–L4 ROM compared with intact in flexion-extension (8.7±2.0 degrees to 12.2±3.2 degrees, p<.05) lateral bending (9.0±2.5 degrees to 12.6±3.2 degrees, p=.09), and axial rotation (3.8±2.7 degrees to 7.8±4.5 degrees p<.05). Pedicle screw fixation decreased ROM compared with intact, resulting in 1.7±0.5 degrees flexion-extension (p<.05), 3.3±1.4 degrees lateral bending (p<.05), and 1.8±0.6 degrees axial rotation (p=.09). TFAS restored intact ROM (p>.05) resulting in 7.9±2.1 degrees flexion-extension, 10.1±3.0 degrees lateral bending, and 4.7±1.6 degrees axial rotation. Fusion significantly increased the normalized ROM at all remaining lumbar segments, whereas TFAS implantation resulted in near-normal distribution of normalized ROM at the implanted and remaining lumbar segments. Flexion and extension stiffness in the high-flexibility zone decreased after facetectomy (p<.05) and increased after simulated fusion (p<.05). TFAS restored quality of motion parameters (load-displacement curves) to intact (p>.05). The quality of motion parameters for the whole lumbar spine mimicked L3–L4 segmental results.ConclusionsTFAS restored range and quality of motion at the operated segment to intact values and restored near-normal motion at the adjacent segments.     

T cell responses to hepatitis B surface antigen are detectable in non-vaccinated individuals

AIM： To evaluate, whether humoral hepatitis-B-vaccine non-responders also fail to mount a T cell response and to compare these results to normal vaccinees. METHODS： Fourty-seven health care employees were enrolled in this study including all available non- responders （n = 13） with an anti-HBsAg titer 〈 10 kU/L and all available low-responders （n = 12） with an anti- HBsAg titer 〈 100 kU/L. Also, 12 consecutive anti-HBsAg negative pre-vaccination subjects were enrolled as well as 10 subjects （＋7 from the vaccinated group） with titers 〉 1000 kU/L as controls. PBMC from all subjects were analyzed by IFN-γ and IL-4 ELISPOT assays for the presence of hepatitis B surface antigen （HBsAg） reactive T cells. RESULTS： Non-responders and low-responders had no or only very limited T cell responses, respectively. Indi- viduals responding to vaccination with the induction of a high anti-HBsAg titer showed a strong T cell response after the third vaccination. Surprisingly, these individuals showed response even before the first vaccination. T cell response to control antigens and mitogens was similar in all groups. CONCLUSION： Our data suggest that there is no gen- eral immune deficiency in non-/low-responders. Thus, we hypothesize that the induction of anti-HBsAg re- sponses by vaccination is significantly dependent on the pre-existing T cell repertoire against the specific antigen rather than the presence of a general T cell defect.     

Endocrinology: Insulin resistant and non-resistant polycystic ovary syndrome represent two clinical and endocrinological subgroups

We studied the clinical and endocrine features of 35 patientswith polycystic ovary syndrome (PCOS) who are either insulinresistant or non-insulin resistant. The occurrence of insulinresistance was determined by measuring insulin and glucose concentrationsfollowing a standard 75 g oral glucose load. All patients wereevaluated by anthropometric measurements: body mass index (BMI),percentage of body fat (BCF) and waist-to-hip ratio (W/H), degreeof hirsutism (Ferriman-Gallwey method) and endocrine profile.Fourteen patients had insulin resistance of unknown origin whereasfour were due to a type A insulin receptor mutation, and 17were non-insulin resistant. The insulin resistant patients weresignificantly more obese (higher BMI P < 0.0001, BCF P <0.002 and W/H ratio P < 0.005) and were more hirsute (P <0.002) than the non-insulin resistant patients. Testosteroneconcentrations were significantly higher in the insulin resistantgroup than in the non-insulin resistant group (2.65 versus 1.37nmol/l; P < 0.027), whereas sex hormone-binding globulinwas lower in insulin resistant patients (30.61 versus 19.48nmol/l; P < 0.02). Non-insulin resistant patients showeda high luteinizing hormone to follicle stimulating hormone ratio,while a normal ratio was found in the insulin resistant subpopulation(2.94 versus 1.34; P < 0.0001). We concluded that PCOS comprisestwo subpopulations, one with insulin resistance of differentaetiologies and the other which has no insulin resistance. Thesetwo groups differ in their anthropometric and endocrine features.The diagnosis of insulin resistance in PCOS can be easily determinedby the insulin response to an oral glucose tolerance test.     

Familial hyperkalemia, hypertension, and hyporeninemia with normal aldosterone levels. A tubular defect in potassium handling

A 52-year-old man had hypertension, persistent hyperkalemia, and hyperchloremic metabolic acidosis; renal and adrenal functions were normal. Four other members of the family have the same findings. The patient's plasma aldosterone (PA) level was within normal range, though plasma renin activity (PRA) was undetectable. The ability to conserve sodium with increased endogenous aldosterone levels, and the inability to increase potassium excretion while exogenous mineralocorticoid (fludrocortisone acetate) was administered, indicated a distal tubular defect in potassium handling. Effective reduction of the hyperkalemia by K+ -Na+ exchange resin also corrected the acidosis and the hyperchloremia, suggesting that hyperkalemia may cause metabolic acidosis.     

How moving objects become animated: the human mirror neuron system assimilates non-biological movement patterns

The so-called mirror neuron system (MNS) responds when humans observe actions performed by a member of their own species. This activity is understood as an internal motor representation of the observed movement pattern. By contrasting meaningless human hand movements with meaningless artificial movements of objects in space, we tested the claim that exclusively movements belonging to the human motor repertoire have direct access to the MNS. Eighteen participants observed video clips of moving hands and objects while the hemodynamic response was recorded with functional magnetic resonance imaging. Second-level analysis of the hemodynamic response revealed substantially overlapping activation patterns for both types of movements including relevant structures of the MNS (bilateral premotor and parietal areas, occipito-temporal junction, postcentral gyrus and the right superior temporal sulcus). This suggests that perceptual processing of moving hands and objects recruits similar and overlapping cortical networks. Direct comparison of the two movement types revealed stronger activations for hand movements mainly in structures of the MNS suggesting an "expertise effect". Overall, our results provide evidence that observing movements not explicitly belonging to the human motor repertoire can activate the human MNS, most likely because an association with a biological movement is evoked.