Suppression of apoptosis in hematopoietic factor-dependent progenitor cell lines by expression of the FAC gene

Fanconi anemia (FA) is a genetically heterogeneous, inherited blood disorder characterized by bone marrow failure, congenital malformations, and a predisposition to leukemias. Because FA cells are hypersensitive to DNA cross-linking agents and have chromosomal instability, FA has been viewed as a disorder of DNA repair. However, the exact cellular defect in FA cells has not been identified. Sequence analysis of the gene defective in group C patients (FAC) has shown no significant homologies to other known genes. The FAC protein has been localized to the cytoplasm, indicating that FAC may either play an indirect role in DNA repair or is involved in a different cellular pathway. Recent evidence has indicated that FA cells may be predisposed to apoptosis, especially after treatment with DNA cross-linking agents. The demonstration that genes can suppress apoptosis has been accomplished by overexpression of such genes in growth factor-dependent cell lines that die by apoptosis after factor withdrawal. Using retroviral-mediated gene transfer, we present evidence that expression of FAC in the hematopoietic factor-dependent progenitor cell lines 32D and MO7e can suppress apoptosis induced by growth factor withdrawal. Flow cytometry and morphologic analysis of propidium iodide stained cells showed significantly lower levels of apoptosis in FAC-retroviral transduced cells after growth factor deprivation. Expression of FAC in both cell lines promoted increased viability rather than proliferation, which is consistent with other apoptosis-inhibiting genes such as Bcl- 2. These findings imply that FAC may act as a mediator of an apoptotic pathway initiated by growth factor withdrawal. Furthermore, the congenital malformations and hematologic abnormalities characterizing FA may be related to an increased predisposition of FA progenitor cells to undergo apoptosis, particularly in the absence of extracellular signals.     

Characterization of a new animal model of chronic hyperprolactinemia

Animal models of chronic prolactin (PRL) excess have included rats bearing transplantable pituitary tumors that have produced other hypophyseal hormones in addition to PRL. We report characterization of a new model, the Buffalo rat implanted with the MMQ tumor, a line developed from the 7315a line. Rats implanted with the MMQ tumor have serum PRL levels that increase with time and correlate with the estimated volume of the subcutaneous tumor. When rats are killed 4 weeks after implantation, serum PRL levels are strikingly higher in tumor-bearing rats compared with controls (females, 2,723 +/- 266 v 192 +/- 46 ng/mL, P less than .0001; males, 1,637 +/- 213 v 99 +/- 11 ng/mL, P less than .0001). Serum PRL levels measured by the Nb2 lymphoma assay were higher than immunoassay measurements in both tumor-bearing and control Buffalo rats. Sephadex chromatography of serum from tumor-bearing rats showed that most of the PRL immunoreactivity co-eluted with 125I-rPRL. Neither serum growth hormone (GH) nor luteinizing hormone (LH) levels were different from controls in tumor-bearing rats. Female MMQ-bearing rats had lower estradiol levels. At death, the wet weights of adrenal glands, kidneys, and gonads were not affected by the presence of tumor. In contrast, tumor-bearing rats had increased spleen weight and histological evidence of white pulp hyperplasia. The Buffalo rat implanted with the PRL-only MMQ tumor is a promising new tool for the study of chronic hyperprolactinemia.     

The Secretion of Endothelin-1 by Microvascular Endothelial Cells from Human Benign Prostatic Hyperplasia is Inhibited by Vascular Endothelial Growth Factor

Prostate growth seems to be influenced by paracrine factors like endothelin-1 (ET-1), originating from the microvascular endothelium. Recently, we reported on the first isolation and primary culture of microvascular endothelial cells (HPEC) derived from tissue of human benign prostatic hyperplasia (BPH). Therefore, direct investigation of growth factor secretion by HPEC is now possible.

BPH tissue was cut into small cubes and gently squeezed after incubation with dispase. HPEC were cultured from the resulting cell suspension after a stepwise selection by use of superparamagnetic beads coated with antibodies against endothelial specific antigens. HPEC were characterized by flow cytometry. After the incubation of HPEC either with vascular endothelial growth factor (VEGF), tumor necrosis factor α (TNF-α), or adenosine triphosphate (ATP), the secretion of ET-1 was measured by ELISA.

HPEC showed a typical endothelial morphology. They were positive for von Willebrand factor and CD31. The ET-1 secretion of HPEC was inhibited by VEGF, but was unaffected by TNF-α or ATP. Furthermore, histochemistry revealed that in vivo microvascular endothelial cells were negative for ET-1. Because of the suppression by the widespread VEGF, it is unlikely that ET-1 from the microvascular endothelium acts as a growth factor in human BPH.     

Hemostatic plug formation in normal and von Willebrand pigs: the effect of the administration of cryoprecipitate and a monoclonal antibody to Willebrand factor

Hemostatic plug (HP) formation was investigated in the ear bleeding time incision in normal and von Willebrand pigs. HP volume was calculated by integrating the areas of serial sections. In normal pigs (n = 11), platelets immediately formed a layer on the surface of the cut channel. Platelet aggregates formed at the ends of transected vessels and gradually enlarged. Finally, all transected vessels were occluded by HP and bleeding stopped. In contrast, large HPs were formed in the incision in von Willebrand's disease (vWD) pigs (n = 4); these HPs did not cover the ends of the transected vessels, which continued to bleed, allowing the formation of large hemostatically ineffective platelet aggregates in the incision. Canals traversed these HPs, and bleeding from the open vessels may have continued through them. After infusion of cryoprecipitate into a vWD pig, the bleeding time shortened, and the morphological findings of the HPs were similar to those of normal pigs. In normal pigs (n = 3) infused with an anti- Willebrand factor monoclonal antibody, which prolonged the bleeding time, a large HP formed in the incision, similar to that observed in the vWD pig. The volume of the normal and vWD HPs increased with time. These in vivo findings suggest that Willebrand factor is involved in the localization of the HP to the damaged vessel and may also play a role in platelet-platelet interaction. A computerized morphometric technique was used for measuring the volume of the hemostatic plugs and the distance of sequential points on the perimeter of the HP from the center of selected bleeding vessels.     

Wound healing effect of methanolic leaf extract of Napoleona vogelii(Family:Lecythidaceae) in rats

Objective:To investigate the wound healing properly of Napoleona vogelii leaf extract in folkloric medicine.Methods:Roth sexes of adult albino rats(n=25) were used in this study and another group(n=30) were subjected to acute toxicity test(LD_(50)) of the plant extract.For the LD_(50),three randomized groups of 5 rats were first treated with 10,100,1 000 mg/kg body weight(bw),orally.This w as followed by a second treatment of 1500,3000,and 5 000 mg/kg bw of the leaf extract with continual monitoring of the animals for mortality or non-mortality.Incision wounds(1.3cm) were created on the skin of five groups of 5 rals using surgical blade under anesthesia.The first group was topically treated with petroleum jelly alone,group 2 was topically applied 400 mg/mL w/v of the reference drug,Neobaein,while group 3-5 were topically treated with 5-50 mg/mL w/v of the plant extract,respectively.Results:The percentage yield of the extract was 49.80%w/w dry matter.The phytochemical analysis revealed several bioactive constituents including glycosides,tannins,alkaloids,perpenoids.saponins,steroids,proteins,and carbohydrates.The LD_(50) was beyond our experimental limit and was not determined.Increased concentrations(5,20,and 50mg/mL w/v) of the extract had significant(ANOVA,P0.05) healing effect on the incision wounds giving rise to 125%-140% while treatmentawith Neobacin resulted in 150% healing effect on the third treatment regimen compared to the control(100%).Conclusions:These data indicate that Napoleona vogelii leaf extract contains potent bioactive compounds containing wound healing activity,substantiating its use as a wound healer in folkloric medicine.     

Hollow screws: a diagnostic tool for intracranial empyema

Background

Subdural (SDE) and epidural empyema (EDE) are life-threatening intracranial infections. They require immediate diagnosis and treatment. However, in some cases, magnet resonance imaging (MRI) is not able to contribute to diagnosis; therefore, surgical exploration is indicated. Hollow screws used for decompression of chronic subdural haematoma (cSDH) are valuable tools for minimally invasive biopsy in awake patients when SDE and EDE are suspected.

Methods

Between 2006 and 2010, eight patients in our department underwent biopsy of a suspected SDE or EDE using hollow screws. In these cases, MRI or computed tomography (CT) were not able to provide sufficient diagnostic security to indicate primary craniotomy. Diagnostic and therapeutic efficacy was evaluated on preoperative and postoperative imaging. The focus was on qualitative parameters, such as contrast enhancement or impaired diffusion on diffusion-weighted images (DWI).

Results

The application of the hollow screw under local anaesthesia permitted an exact diagnosis in all cases. In one case, the suspected diagnosis of cSDH could be refuted by diagnostic puncture. In four cases of uncertain diagnosis, the application of the hollow screw revealed a cSDH. Seven of eight patients previously received neurosurgical treatment; three of those cases were SDE or EDE and four were cSDH. Cases of SDE and EDE needed further craniotomy after diagnostic puncture, whereas patients with cSDH were sufficiently treated by hollow screws.

Conclusions

Given their comparably wide diameter, hollow screws allow a sufficient sample size and, therefore, lead to precise diagnosis of SDE and EDE without significant operative risks or strains for the patient.     

Kardiales CT und Ablation von Vorhofflimmern

Both cardiac computed tomography (CT) and interventional electrophysiology (EP) have evolved considerably in recent years. Technical improvements in CT have significantly reduced the radiation dose in cardiac applications. This imaging technology plays an important role in preprocedural planning and guidance of the procedures in many EP centers worldwide. Furthermore, CT is the imaging modality of choice to diagnose relevant complications in ablation of atrial fibrillation, e.g. pulmonary vein stenosis or atrioesophageal fistula. In anatomically driven ablation procedures, such as balloon-based procedures in atrial fibrillation, detailed analysis of the relevant cardiac structures is absolutely crucial not only to reduce radiation exposure and procedure times but also to improve ablation success and to reduce the occurrence of complications. Current software applications enable 3-dimensional reconstruction of cardiac images and the integration into electroanatomical navigation systems. This article reviews the available evidence in this field and highlights recent developments in image guidance for ablation of atrial fibrillation.