Subcellular localization of transforming growth factor-alpha in human eosinophil granulocytes

Eosinophils are involved in the inflammatory response seen in allergy and helminthic infestations. Eosinophils synthesize transforming growth factor-alpha (TGF-alpha), which may play a role in the development of the characteristic fibrosis seen in longstanding high eosinophilia. Using immunoelectron microscopic techniques, eosinophils from peripheral blood of healthy individuals and from one patient with high eosinophilia showed presence TGF-alpha in matrix of the specific crystalloid-containing granules. In cryosections, TGF-alpha was also visualized in a vesicular compartment of the cytoplasm. In double- labeling experiments, the TGF-alpha of this latter compartment did not colocalize with CD63, a marker for lysosomes, nor with albumin of secretory vesicles. In extracts from eosinophils, obtained from healthy donors, immunoreactive TGF-alpha could be detected by enzyme-linked immunosorbent assay-technique. In addition, sera from two patients with high eosinophilia showed TGF-alpha concentrations of 1.5 ng/mL and 164 pg/mL, respectively, whereas TGF-alpha could not be detected in serum from healthy controls. In conclusion, TGF-alpha is present in the specific granules, and in an additional vesicular compartment of the cytoplasm of eosinophils.     

Transplantation potential of hematopoietic cells released into the circulation during routine chemotherapy for non-Hodgkin's lymphoma [see comments]

Primitive hematopoietic cells released into the peripheral blood (PB) were studied in 50 patients with high-grade non-Hodgkin's lymphoma enrolled in a phase III trial of intensive weekly chemotherapy (VAPEC- B) alone or with granulocyte colony-stimulating factor (G-CSF). Mononuclear cells numbers were monitored and their in vitro growth potential assessed in clonogenic progenitor cell assays and in long- term culture. Total colony-forming cells (granulocyte-macrophage [GM], burst-forming unit, erythroid [BFU-E], Mix-CFC) were increased 40-fold (median) over baseline with chemotherapy alone and 106-fold with chemotherapy and G-CSF after the final dose. CD34+ cells were increased to a median of 4%, equivalent to that in normal bone marrow (BM) controls. Circulating colony-forming cell levels were maximal when the recovering total white blood cell (WBC) count reached 5 to 10 x 10(9)/L. The timing of the maximum was reproducible in individual patients. Therefore the WBC count can be used as a guide to the timing of leukapheresis. PB cells from normal controls' and patients' prechemotherapy were unable to sustain hemopoiesis in two-stage long- term cultures. In contrast, PB cells collected from patients primed with chemotherapy alone or chemotherapy with G-CSF at the time of predicted maximal colony-forming cell release were able to generate and sustain hematopoiesis in long-term cultures at a level comparable or superior to normal BM. These findings indicate that the use of G-CSF after routine outpatient chemotherapy stimulates maximal release of primitive hemopoietic cells into the circulation, including colony- forming cells and long-term culture-initiating cells. Their numbers are comparable with those in normal BM and are such that a single leukapheresis will usually yield enough cells for hemopoietic reconstitution after myeloablative chemotherapy.     

Effects of Intravenous Injection of Adipose‐Derived Stem Cells in a Rat Model of Radiation Therapy‐Induced Erectile Dysfunction

IntroductionRadiation therapy (RT) for prostate cancer is frequently associated with posttreatment erectile dysfunction (ED).AimTo investigate whether injection of adipose‐derived stem cells (ADSCs) can ameliorate RT‐associated ED.MethodsThirty male rats were divided into three groups. The control + phosphate‐buffered saline (PBS) group received tail‐vein injection of PBS. The radiation + PBS group received radiation over the prostate and tail‐vein injection of PBS. The radiation + ADSC group received radiation over the prostate and tail‐vein injection of ADSCs, which were labeled with 5‐ethynyl‐2‐deoxyuridine (EdU). Seventeen weeks later, erectile function was evaluated by intracavernous pressure (ICP) in response to electrostimulation of cavernous nerves (CNs). Penile tissue and major pelvic ganglia (MPG) were examined by immunofluorescence (IF) and EdU staining.Main Outcome MeasuresErectile function was measured by ICP. Protein expression was examined by IF, followed by image analysis and quantification.ResultsRadiation over the prostate caused a significant decrease in erectile function and in the expression of neuronal nitric oxide synthase (nNOS) in penis and MPG. Cavernous smooth muscle (CSM) but not endothelial content was also reduced. Injection of ADSCs significantly restored erectile function, nNOS expression, and CSM content in the irradiated rats. EdU‐positive cells were visible in MPG.ConclusionsRadiation appears to cause ED via CN injury. ADSC injection can restore erectile function via CN regeneration. Qiu X, Villalta J, Ferretti L, Fandel TM, Albersen M, Lin G, Dai Y, Lue TF, and Lin C‐S. Effects of intravenous injection of adipose‐derived stem cells in a rat model of radiation therapy‐induced erectile dysfunction. J Sex Med 2012;9:1851–1858.     

Injections of Adipose Tissue-Derived Stem Cells and Stem Cell Lysate Improve Recovery of Erectile Function in a Rat Model of Cavernous Nerve Injury

IntroductionErectile dysfunction (ED) remains a major complication after radical prostatectomy. The use of adipose tissue-derived stem cells (ADSCs) has shown promising results for the treatment of ED. However, the mechanisms of action for stem cell therapy remain controversial, with increasing evidence pointing to paracrine pathways.AimTo determine the effects and to identify the mechanism of action of ADSC and ADSC-derived lysate in a rat model of cavernous nerve (CN) crush injury.MethodsThirty-two male Sprague-Dawley rats were randomly divided into four equal groups: one group underwent sham operation, while three groups underwent bilateral CN crush. Crush-injury groups were treated at the time of injury with intracavernous injection of ADSC, lysate, or vehicle only (injured controls). Erectile function was assessed by CN electrostimulation at 4 weeks. Penile tissue was collected for histology.Main Outcome MeasuresIntracavernous pressure increase upon CN stimulation; neuronal nitric oxide synthase (nNOS) content in the dorsal penile nerve; smooth muscle content, collagen content, and number of apoptotic cells in the corpus cavernosum.ResultsBoth ADSC and lysate treatments resulted in significant recovery of erectile function, as compared with vehicle treatment. nNOS content was preserved in both the ADSC and lysate group, with significantly higher expression compared with vehicle-treated animals. There was significantly less fibrosis and a significant preservation of smooth muscle content in the ADSC and lysate groups compared with injured controls. The observed functional improvement after lysate injection supports the hypothesis that ADSCs act through release of intracellular preformed substances or by active secretion of certain biomolecules. The underlying mechanism of recovery appears to involve neuron preservation and cytoprotection by inhibition of apoptosis.ConclusionsPenile injection of both ADSC and ADSC-derived lysate can improve recovery of erectile function in a rat model of neurogenic ED. Albersen M, Fandel TM, Lin G, Wang G, Banie L, Lin CS, and Lue TF. Injections of adipose tissue-derived stem cells and stem cell lysate improve recovery of erectile function in a rat model of cavernous nerve injury.     

"Stem cell" origin of the hematopoietic defect in dyskeratosis congenita

We have used the long-term bone marrow culture (LTBMC) system to analyze hematopoiesis in three patients with dyskeratosis congenita (DC), two of whom had aplastic anemia, and the third had a normal blood count (apart from mild macrocytosis) and normal BM cellularity. Hematopoiesis was severely defective in all three patients, as measured by a low incidence of colony-forming cells and a low level of hematopoiesis in LTBMC. The function of the marrow stroma was normal in its ability to support the growth of hematopoietic progenitors from normal marrows seeded onto them in all three cases, but the generation of hematopoietic progenitors from patients marrow cells inoculated onto normal stromas was reduced, thus suggesting the defect to be of stem cell origin. The parents and unaffected brother of one of the families have also been studied in LTBMC and all showed normal hematopoietic and stromal cell function. From this study we speculate that there are some similarities between DC and the defect in the W/Wv mouse.