Experimental calibration of a two-stage prism-grating system for measuring cell velocity

A new optical system for the measurement of the erythrocyte velocity in microvessels is extensively applied for the first time. It is based on the projection of the erythrocyte image onto two photodiodes through a prism grating. The differential signal of the photodiodes is proportional to the velocity. The system is comparable to those of M. Anliker, M. Casty, P. Friedli, R. Kubli, and H. Keller ((1977). Noninvasive measurement of blood flow. In “Cardiovascular Flow Dynamics and Measurement” (N. H. Hwang and N. Normann, eds.), pp. 43–88. Univ. Park Press, Baltimore) and D. W. Slaaf, J. P. S. M. Rood, G. J. Tangelder, and T. Arts ((1979). Microvasc. Res.17, S173). It measures an instantaneous velocity. The system is calibrated by a velocity which is constant over the total measuring field and by a known flow rate and profile. Concerning applicability and results the device is compared to the two-slit methods, the laser-Doppler-anemometry and high-speed cinematography.     

Comparative Efficacies and Tolerabilities of Intravenous Azithromycin Plus Ceftriaxone and Intravenous Levofloxacin with Step-Down Oral Therapy for Hospitalized Patients with Moderate to Severe Community-Acquired Pneumonia

Objective: To compare the efficacy and tolerability of ceftriaxone plus azithromycin with those of levofloxacin in the treatment of hospitalized patients with moderate to severe community-acquired pneumonia (CAP). Design: Randomized, open-label multicenter trial with 1 : 1 treatment allocation in an inpatient setting. Patients: 212 male or female inpatients with a clinical diagnosis of CAP were included in the study. In each treatment group >50% of patients had a pneumonia severity index of IV or V. Interventions: Open-label treatment with either intravenous (IV) ceftriaxone 1g and IV azithromycin 500mg daily or IV levofloxacin 500mg daily. Patients who improved clinically were switched to oral follow-on therapy with either azithromycin 500 mg/day or levofloxacin 500 mg/day. At the clinician’s discretion, oral cefuroxime axetil was added to the treatment regimen of patients who received oral azithromycin if a macrolide resistant pneumococcal isolate was documented. Results: Overall, both study treatments were well tolerated. Favorable clinical outcomes in clinically evaluable patients were demonstrated in 91.5% of patients treated with ceftriaxone plus azithromycin and 89.3% (95% CI ?7.1%, 11.4%) of patients treated with levofloxacin at the end of therapy visit and in 89.2% and 85.1% (95% CI ?6.7%, 14.8%) patients, respectively, at the end of study visit. Bacteriological eradication rates for both treatments were equivalent with the exception of Streptococcus pneumoniae; 44% of isolates were eradicated with levofloxacin compared with 100% of isolates with ceftriaxone plus azithromycin. Conclusions: As acknowledged by international CAP treatment guidelines, the combination of a third-generation cephalosporin and a macrolide is at least as efficacious as monotherapy with a fluoroquinolone with enhanced anti-pneumococcal activity, for hospitalized patients with moderate to severe CAP. Combined medication with a macrolide and third-generation cephalosporin may be preferred over fluoroquinolones as first-line therapy of hospitalized patients with CAP to minimize the development of multiresistant nosocomial Gram-negative bacilli.     

Recent developments in antibiotic treatment

Antibiotic resistance of gram-positive and gram-negative bacteria remains a major challenge for clinicians treating HAP. Since the recent release of linezolid and QD, treatment options for resistant gram-positive bacteria have improved. The development of new substances continues and it is hoped that some of them will be available soon. Investigation has centered on gram-positive bacteria, although multiresistant gram-negative pathogens, such as A haumanii, S maltophilia, and resistant P aeruginosa, are of major clinical relevance. New treatment options are unfortunately not in sight. No antibiotic, however, is a miraculous magic wand against resistant bacteria. The bugs are smart; they have been on this world far longer than humans. Regardless of how innovative the mechanism of action of new substances is, resistance will emerge. The solution is certainly not a nihilistic approach leading to a fearful restriction in the use of new substances. No antibiotic, regardless of its potency, can free the clinician from keeping the difficult balance between individual undertreatment and general overtreatment.     

Peripheral blood progenitor cells mobilized by chemotherapy plus granulocyte-colony stimulating factor accelerate both neutrophil and platelet recovery after high-dose VP16, ifosfamide and cisplatin

Summary. We report on the chemotherapy plus granulocyte colony-stimulating factor (G-CSF) induced mobilization of peripheral blood progenitor cells (PBPCs) and their impact on haematopoietic recovery following high-dose chemotherapy. Twenty-four patients with advanced solid tumours or lymphomas received standard-dose chemotherapy with VP16, ifosfamide and cisplatin (VIP) followed by filgrastim (G-CSF; 5 μg/kg s.c. daily for 14 d) for the prevention of chemotherapy induced neutropenia and for the simultaneous mobilization of PBPCs. Maximal numbers of progenitors of different lineages were reached at day 11 (range 9–14) after VIP chemotherapy. A median of 0·415 × 10⁹/1 CD34⁺ cells (range 0·11–1·98), 9000 CFU-GM/ml (range 2800–17700). 3500 BFU-E/ml (range 400–10800) and 200 CFU-GEMM/ml (range 0–4400) were recruited. One single apheresis yielded a median of 1·6 × 10⁸ mononuclear cells/kg (range 0·2–5·4) or 5·4 × 10⁶ CD34⁺ cells/kg body weight (range 0·2–24·2). Fourteen patients who showed at least a partial remission after two cycles of the standard-dose chemotherapy regimen were subjected to high-dose VIP chemotherapy (cumulative doses of 1500 mg/m² VP16, 12 g/m² ifosfamide and 150 mg/m² cisplatin) with or without PBPC support. The first six patients were treated with growth factors only (IL-3/GM-CSF) and did not receive PBPCs, whereas the following eight patients were supported with PBPCs in addition to IL-3 and GM-CSF. Neutrophil recovery as well as platelet recovery were significantly faster in patients receiving PBPCs with a median of 6·5 d below 0·1 × 10⁹ neutrophils/1 and 3 d below 20 × 10⁹ platelets/1 as compared to 10·5 d and 8 d in control patients receiving growth factors only. The accelerated platelet recovery in patients supported with PBPCs might be explained—in the absence of detectable colony-forming units megakaryocyte—by the presence of glycoprotein IIb/IIIa⁺, non-proliferating endomitotic megakaryocytic precursor cells within G-CSF mobilized PBPCs. Our data demonstrate that chemotherapy plus G-CSF mobilized PBPCs accelerate both neutrophil and platelet recovery after high-dose VIP chemotherapy in patients with solid tumours or lymphomas.     

Value of comparative genomic hybridization and fluorescence in situ hybridization for molecular diagnostics in multiple myeloma

Chromosomal abnormalities, such as 13q deletions, are emerging as important prognostic factors in multiple myeloma. Fluorescence in situ hybridization (FISH) using specific DNA probes is the technique most widely used for the determination of genomic aberrations in this disease. The utility of comparative genomic hybridization (CGH) for molecular diagnostics in plasma cell malignancies has not been systematically analysed. We investigated tumour samples of patients with multiple myeloma (n = 43) or plasma cell leukaemia (n = 3) using CGH and FISH with five DNA probes localized to chromosome bands 1p22, 6q21, 11q22-q23, 13q14 and 17p13. By CGH, the most frequent genomic changes were gains on chromosomes 1q, 9q and 11q, as well as losses on chromosomes 13q, 6q, Xp and Xq. By FISH, trisomy 11q was identified at a similar frequency to the 13q deletion (42%). Compared with FISH data, the sensitivity of CGH was 80.7% and the specificity was 97.5%. Thirty-two aberrations found by FISH were not identified by CGH, mostly as a result of the proportion of cells carrying the respective aberrations, or because of the limited spatial resolution of CGH. Our data indicate that, for clinical molecular diagnostics in multiple myeloma, FISH with a disease-specific DNA probe set is superior to CGH analysis.     

Up-regulation of endothelial stretch-activated cation channels by fluid shear stress.

OBJECTIVE: Stretch-activated cation channels (SAC) have been suggested to act as endothelial mechanosensors for hemodynamic forces. Ca(2+) influx through SAC could induce an intracellular Ca(2+) signal stimulating Ca(2+)-dependent synthesis of vasodilators like NO, prostacyclin, or EDHF. In the present study we tested whether laminar shear stress (LSS) regulates SAC function. METHODS: Electrophysiological properties of SAC were investigated in human umbilical vein endothelial cells (HUVEC) subjected to defined levels of LSS in a flow-cone apparatus. RESULTS: In HUVEC, we identified a Ca(2+) permeable SAC that was activated by membrane stretch. Single-channel current densities of SAC in cell-attached patches were significantly increased in HUVEC exposed to an LSS of 5 dyn/cm(2) for 4 h (1.15+/-0.17 SAC/patch) compared to HUVEC kept in stationary culture (0.46+/-0.07 SAC/patch). Exposure of HUVEC to a higher LSS of 15 dyn/cm(2) for 4 h induced similar up-regulation of SAC (1.27+/-0.21 SAC/patch). After 24 h exposure to LSS of 15 dyn/cm(2), single-channel current densities of SAC remained up-regulated (1.07+/-0.18 SAC/patch) compared to controls. In addition, stretch-sensitivity of SAC (channel activity NP(o) at -30 mmHg) significantly increased after 2 h of exposure to LSS of 5 and 15 dyn/cm(2) and remained up-regulated after 24 h. Inhibition of protein kinases and tyrosine kinases by H7 and genistein, respectively, prevented LSS-induced alteration of SAC function. CONCLUSION: Single-channel current density and mechanosensitivity of SAC in HUVEC is up-regulated by LSS. Up-regulation of SAC function leads to enhanced mechanosensitive Ca(2+) influx, and represents a novel adaptive mechanism of the endothelium in the presence of altered hemodynamic forces.     

Impaired survival of peripheral T cells, disrupted NK/NKT cell development, and liver failure in mice lacking Gimap5

The loss of Gimap5 (GTPase of the immune-associated protein 5) gene function is the underlying cause of lymphopenia and autoimmune diabetes in the BioBreeding (BB) rat. The in vivo function of murine gimap5 is largely unknown. We show that selective gene ablation of the mouse gimap5 gene impairs the final intrathymic maturation of CD8 and CD4 T cells and compromises the survival of postthymic CD4 and CD8 cells, replicating findings in the BB rat model. In addition, gimap5 deficiency imposes a block of natural killer (NK)- and NKT-cell differentiation. Development of NK/NKT cells is restored on transfer of gimap5(-/-) bone marrow into a wild-type environment. Mice lacking gimap5 have a median survival of 15 weeks, exhibit chronic hepatic hematopoiesis, and in later stages show pronounced hepatocyte apoptosis, leading to liver failure. This pathology persists in a Rag2-deficient background in the absence of mature B, T, or NK cells and cannot be adoptively transferred by transplanting gimap5(-/-) bone marrow into wild-type recipients. We conclude that mouse gimap5 is necessary for the survival of peripheral T cells, NK/NKT-cell development, and the maintenance of normal liver function. These functions involve cell-intrinsic as well as cell-extrinsic mechanisms.