Exclusion of dysfunctional mitochondria from Balbiani body during early oogenesis of Thermobia.

Oocytes of many invertebrate and vertebrate species contain a characteristic organelle complex known as the Balbiani body (Bb). Until now, three principal functions have been ascribed to this complex: delivery of germ cell determinants and localized RNAs to the vegetal cortex/posterior pole of the oocyte, transport of the mitochondria towards the germ plasm, and participation in the formation of lipid droplets. Here, we present the results of a computer-aided 3D reconstruction of the Bb in the growing oocytes of an insect, Thermobia domestica. Our analyses have shown that, in Thermobia, the central part of each fully developed Bb comprises a single intricate mitochondrial network. This "core" network is surrounded by several isolated bean-shaped mitochondrial units that display lowered membrane potential and clear signs of degeneration. In light of the above results and recent theoretical models of mitochondrial quality control, the role of the Bb is discussed. We suggest that, in addition to the aforementioned functions, the Bb is implicated in the selective elimination of dysfunctional mitochondria during oogenesis.

The use of Yttrium-90 Ibritumomab Tiuxetan ((90)Y-IT) as a consolidation therapy in high-risk patients with diffuse large B-cell lymphoma ineligible for autologous stem-cell transplantation.

AIM OF THE STUDY: To evaluate the efficacy and safety of Yttrium-90 Ibritumomab Tiuxetan ((90)Y-IT) as a consolidation therapy in the management of DLBCL. MATERIAL AND METHODS: Patients with primary refractory or high-risk DLBCL (n = 18), ineligible for autologous stem-cell transplantation, were included in a retrospective study performed at three centers by the Polish Lymphoma Research Group (PLRG). All patients (mean age 61, range 35-82) either didn't achieve a complete response or didn't complete the scheduled therapy due to its complications. Response rates (CR, PR, SD, PD) according to Cheson criteria, overall survival (OS), progression-free survival (PFS) and adverse effects of radioimmunotherapy were analyzed. RESULTS: Consolidation radioimmunotherapy increased the CR rate from 38% (n = 7) to 82% (n = 15). One patient remained in PR, one patient remained in SD, while one patient remained in PD. During a median follow-up of five years, 11 patients (62%) were alive with no recurrence, 4 patients (22%) were alive with relapse while 3 patients (16%) died. There was no statistically significant difference in PFS between those in CR and those in PR before (90)Y-IT. CONCLUSIONS: Radioimmunotherapy is an effective consolidation therapy for high risk/refractory DLBCL patients and worthy of further investigation in prospective trials.

[Analysis of the prevalence of HBV markers relevant to the risk of reactivation of infection in patients with hematologic diseases]. / Analiza wystepowania markerów wirusa HBV istotnych dla ryzyka reaktywacji zakazenia u osób z chorobami hematologicznymi.

The study assessed the incidence of HBV markers (HBsAg, anti-HBc, anti-HBs) important for determination of the risk of reactivation of infection, with particular interest of occult infection (presence of HBV DNA in the absence of HBsAg) in patients treated at the Institute of Hematology and Transfusion Medicine. Anti-HBc frequency was correlated with the age and sex of patients. HBsAg was detected in 16/468 examined patients, 98/468 (21%) were anti-HBc positive. HBV DNA was detected in 41/98 anti-HBc positives; in 13 simultaneously with HBsAg. 28 patients had occult HBV infection (HBV DNA+/HBsAg). Antibody to HBsAg was detected in 163/430 (38%) patients, 81 out of them on protective level (> 100 IU/l). It was shown that occult HBV infection occurs in approximately 6% of patients. In most of them the protective levels of anti-HBs are detected.

Hematopoietic stem cell mobilization with the reversible CXCR4 receptor inhibitor plerixafor (AMD3100)-Polish compassionate use experience.

Recent developments in the field of targeted therapy have led to the discovery of a new drug, plerixafor, that is a specific inhibitor of the CXCR4 receptor. Plerixafor acts in concert with granulocyte colony-stimulating factor (G-CSF) to increase the number of stem cells circulating in the peripheral blood (PB). Therefore, it has been applied in the field of hematopoietic stem cell mobilization. We analyzed retrospectively data regarding stem cell mobilization with plerixafor in a cohort of 61 patients suffering from multiple myeloma (N = 23), non-Hodgkin's lymphoma (N = 20), or Hodgkin's lymphoma (N = 18). At least one previous mobilization attempt had failed in 83.6% of these patients, whereas 16.4% were predicted to be poor mobilizers. The median number of CD34+ cells in the PB after the first administration of plerixafor was 22/µL (range of 0-121). In total, 85.2% of the patients proceeded to cell collection, and a median of two (range of 0-4) aphereses were performed. A minimum of 2.0 × 10(6) CD34+ cells per kilogram of the patient's body weight (cells/kg b.w.) was collected from 65.6% of patients, and the median number of cells collected was 2.67 × 10(6) CD34+ cells/kg b.w. (0-8.0). Of the patients, 55.7% had already undergone autologous stem cell transplantation, and the median time to neutrophil and platelet reconstitution was 12 and 14 days, respectively. Cases of late graft failure were not observed. We identified the diagnosis of non-Hodgkin's lymphoma and previous radiotherapy as independent factors that contributed to failure of mobilization. The current report demonstrates the satisfactory efficacy of plerixafor plus G-CSF for stem cell mobilization in heavily pre-treated poor or predicted poor mobilizers.

A very simple mode of follicular cell diversification in Euborellia fulviceps (Dermaptera, Anisolabididae) involves actively migrating cells.

The ovaries of Euborellia fulviceps are composed of five elongated ovarioles of meroistic-polytrophic type. The individual ovariole has three discernible regions: the terminal filament, germarium, and vitellarium. The terminal filament is a stalk of flattened, disc-shaped somatic cells. In the germarium, germline cells in subsequent stages of differentiation are located, and the vitellarium comprises numerous ovarian follicles arranged linearly. The individual ovarian follicles within the vitellarium are separated by prominent interfollicular stalks. The follicles are composed by two germline cells only: an oocyte and a single, polyploid nurse cell, which are surrounded by a monolayer of somatic follicular cells (FCs). During subsequent stages of oogenesis, initially uniform follicular epithelium begins to diversify into morphologically and physiologically distinct subpopulations. In E. fulviceps, the FC diversification mode is rather simple and leads to the formation of only three different FC subpopulations: (1) cuboidal FCs covering the oocyte, (2) stretched FCs surrounding the nurse cell and (3) FCs actively migrating between oocyte and a nurse cell. We found that FCs from the latter subpopulation send long and thin filopodium-like and microtubule-rich processes penetrating between the oocyte and nurse cell membranes. This suggests that, in E. fulviceps, cells from at least one FCs subpopulation show the ability to change position within an ovarian follicle by means of active migration.

Structure of ovaries and oogenesis in dermapterans. II. The nurse cells, nuage aggregates and sponge bodies.

In three studied dermapteran species, Doru lineare, Opisthocosmia silvestris and Forficula auricularia, ovarian follicles are composed of two cells only, the oocyte and a single nurse cell. The nuclei of the nurse cells are large, ameboid and contain highly active nucleoli. RER elements, ribosomes, mitochondria and electron-dense aggregations of nuage material are present in the cytoplasm. Immunolocalization analysis revealed that in earwigs the nuage does not contain snRNAs. In one of the studied species, Doru lineare, apart from "canonical" nuage aggregations, characteristic RER/nuage complexes were found. These structures are morphologically similar to the sponge bodies present in the cytoplasm of the Drosophila germline cells. We suggest that RER/nuage complexes in Doru, as sponge bodies in Drosophila, are implicated in mRNA translocation.

Diversification of follicular cells in the ovaries of the horse fly Haematopota italica (Diptera: Tabanidae). Similarities and differences with the Drosophila model system.

In insect ovaries, germ line cells are surrounded by somatic cells that initially form a uniform follicular epithelium. The subsequent diversification of the follicular cells into several subpopulations enables specification of distinct structures in different regions of complex eggshells. It also influences the patterning of the future embryo. These processes have been extensively studied at both the cellular and molecular levels using the Drosophila ovary as a model system. It is not clear however, to what extent the Drosophila model of the follicular epithelium patterning is universal for the entire Diptera group. Here, we analyze the diversification of the follicular cells in a distant Drosophila relative, the horse fly, Haematopota italica. We found that in this species, there are 6 recognizably different follicular cell subpopulations within the previtellogenic ovarian follicles. Ultrastructural analysis of the follicular epithelium revealed two morphologically distinct clusters of follicular cells residing at the anterior and posterior poles of the follicles. Each cluster consists of 2-3 polar cells located centrally and surrounded by several outer cells called border cells (at the anterior pole) or border-like cells (at the posterior pole). During previtellogenesis, the clusters lose the initial symmetry as their cells differentiate and develop conspicuous cytoplasmic projections comprising cytoskeletal elements. Ultimately, the follicular cells of the anterior and posterior clusters become morphologically different and, as we suggest, participate in different processes during oogenesis and formation of the eggshell in H. italica.

[Replication of HCV in bone marrow of patients with haematological disorders]. / Replikacja wirusa zapalenia watroby typu C (HCV) w szpiku kostnym pacjentów z zaburzeniami hematologicznymi.

UNLABELLED: The aim of the study was to evaluate the presence of HCV replication in bone marrow cells derived from patients displaying hematological disorders. We analysed serum, peripheral blood mononuclear cells (PBMC) and bone marrow samples obtained from 27 patients displaying the following dysfunctions: lymphoma, trombocytopenia, haemophilia, pancytopenia and acute myeloid leukemia (AML). The presence of HCV-RNA in samples was detected by RT-PCR. All the serum samples were HCV-RNA positive as well as 9 out of 27 (33%) PBMC and 17 out of 27 (63%) of bone marrow samples. Independently to the disorder type, the co-presence of HCV-RNA in serum and bone marrow with the simultaneous absence of the viral genetic material in PBMC was detected in 5 (18.5%) of patients. This result suggests that bone marrow is a site of active viral replication. To check whether a viral replication generates any mutations, an SSCP analysis of the 5'UTR viral region was performed. The difference in the viral sequence derived from serum, PBMC and bone marrow was detected in one case. This result may indicate the occurrence of mutation process during the viral replication in bone marrow. An immunohistochemical analysis of bone marrow smears showed the presence of HCV antigens. CONCLUSION: bone marrow cells of patients displaying hematological disorders represent a putative site of extrahepatic HCV replication.

Consolidation with 90Y ibritumomab tiuxetan radioimmunotherapy in mantle cell lymphoma patients ineligible for high dose therapy: results of the phase II multicentre Polish Lymphoma Research Group trial, after 8-year long follow-up.

Polish Lymphoma Research Group performed a phase-II trial to test whether 90Y ibritumomab tiuxetan radioimmunotherapy (Y90) may constitute an alternative consolidation for mantle cell lymphoma patients unfit for high-dose therapy. Forty-six patients were consolidated with Y90 following response to the 1st (n = 34) or 2nd line (n = 12) (immuno)chemotherapy. Majority of the patients had advanced disease (stage IV and presence of B-symptoms in 85% and 70%, respectively) and high MIPI (5.8, range 4-7). Consolidation with Y90 increased the complete remission (CR) rate obtained by the 1st line therapy from 41% to 91% and allowed for median PFS of 3.3 and OS of 6.5 years. In the first relapse, CR rate increased from 16% to 75%, while median PFS and OS totaled 2.2 and 6.5 years, respectively. At 8 years, 30% of patients, consolidated in the 1st line CR were alive, without relapse. Toxicity associated with Y90 is manageable, more severe after fludarabine-based regimens.

[Antiphospholipid antibodies with HCV infection. Innocent proteins or risk factor?]. / Przeciwciala antyfosfolipidowe w zakazeniu HCV. Niewinne bialka, czy czynnik ryzyka?

Infection with hepatitis C virus (HCV) may be associated with a wide spectrum of immunological abnormalities. HCV tends to induce nonspecific autoimmune reactions, as demonstrated by the high prevalence of various autoantibodies, including antiphospholipid antibodies (aPL). The aPL antibodies (lupus anticoagulant and anticardiolipin antibodies) are a heterogeneous family of immunoglobulins reactive with complexes of phospholipids and plasma proteins (cofactors). The most important of these protein cofactors are beta2-glycoprotein (beta2-GPI) and prothrombin. The antiphospholipid syndrome (APS) is an autoimmune disorder characterized by arterial or venous thrombosis, recurrent fetal losses in association with the presence of antiphospholipid (aPL) antibodies. Increased prevalence of aPL antibodies in several bacterial, parasitic, and viral infections have been reported. Most of the published data agree that anticardiolipin antibodies are frequently found in patients with chronic HCV infection, but they do not appear to be of clinical importance. Some studies, however, have found an increased incidence of thrombotic disorders in patients with chronic hepatitis C virus (HCV) who manifest aPL positivity. More prospective, long-term studies are required in order to address whether HCV is involved or not in the etiopathogenesis of APS.

Meiosis, Balbiani body and early asymmetry of Thermobia oocyte.

The meiotic division guarantees maintenance of a genetic diversity; it consists of several stages, with prophase I being the longest and the most complex. We decided to follow the course of initial stages of meiotic division in ovaries of Thermobia domestica using modified techniques of squash preparations, semithin sections, and electron microscopy. We show that germaria contain numerous germline cells that can be classified into three categories: cystoblasts, meiotic oocytes, and growing previtellogenic oocytes. The cystoblasts are located most apically. The meiotic oocytes occupy the middle part of the germarium, and the previtellogenic oocytes can be found in the most basal part, near the vitellarium. Analyses of the semithin sections and squash preparations show that post leptotene meiotic chromosomes gather in one region of the nucleoplasm where they form the so-called bouquet. The telomeres of the bouquet chromosomes are attached to a relatively small area (segment) of the nuclear envelope. Next to this envelope segment, the nucleolar organizers are also located. We show that in concert to sequential changes inside the oocyte nuclei, rearrangement of organelles within the ooplasm (oocyte cytoplasm) takes place. This leads to the formation of the Balbiani body and consequent asymmetry of the ooplasm. These early nuclear and cytoplasmic asymmetries, however, are transient. During diplotene, the chromosome bouquet disappears, while the Balbiani body gradually disperses throughout the ooplasm. Finally, our observations indicate the presence of lampbrush chromosomes in the nuclei of previtellogenic oocytes. In the close vicinity to lampbrush chromosomes, characteristic spherical nuclear bodies are present.

[Isotype switching of produced immunoglobulins in multiple myeloma]. / Przelaczanie klasy produkowanych immunoglobulin w szpiczaku mnogim.

Multiple myeloma is usually characterized by production of a single serum monoclonal protein of constant isotype and light chain restriction. Isotype switching in human lymphomas appears to be an uncommon event. It could be due to altered para-protein production by the malignant plasma cell clone, or oligoclonal Ig production during recovery of B-cell function after chemotherapy. We describe a case of clonal heavy chain isotype switching in a patient with lymphoplasmocytic lymphoma.

Morphology and ultrastructure of the germarium in panoistic ovarioles of a basal "apterygotous" insect, Thermobia domestica.

It has been shown that in Drosophila the germline stem cells (GSCs), similar to the germline and non-germline stem cells of other species, develop and function in specialized microenvironments formed by somatic cells, referred to as the niches. In the fruit fly ovaries, the female GSCs divide asymmetrically to produce new GSCs and the progenitor cells, the cystoblasts (Cbs). Each Cb then divides to generate the cyst composed of 16 interconnected sibling cells, the cystocytes. After cyst formation, specific molecules are transferred to one of the cystocytes which differentiates into the oocyte, whereas the other 15 cystocytes become the nurse cells. We have studied morphology and ultrastructure of the germaria in the ovarioles (ovaries) of a basal "apterygotous" insect, the firebrat (Thermobia domestica). Our analyses have revealed that in this insect, putative GSCs are present along the anterior apex of the germarium. These cells are separated from each other and from the basement lamina covering the ovariole by characteristic somatic cells, termed the apical somatic cells (ASCs), or their elongated processes. We believe that all the ASCs of a given ovariole constitute a "dispersed" niche in which putative GSCs are anchored. Our analyses have additionally shown that in Thermobia, both the Cbs and young (meiotic) oocytes are always individual and never form syncytial cysts. These findings indicate that in certain basal insects the syncytial phase of oogenesis has been eliminated during evolution. Finally, we show that in the early meiotic oocytes of Thermobia, during the so-called bouquet stage, prominent Balbiani bodies (Bbs) are formed. Analysis of serial micrographs indicates that the Bbs invariably arise next to the segment of the nuclear envelope to which the telomeres of the bouquet chromosomes are attached. We suggest, in the light of these data, that the localization of the Bb together with the polar attachment of the bouquet chromosomes play a crucial role in the early asymmetrization of Thermobia oocytes.

Embryos of the viviparous dermapteran, Arixenia esau develop sequentially in two compartments: terminal ovarian follicles and the uterus.

Three main reproductive strategies have been described among insects: most common oviparity, ovoviviparity and viviparity. In the latter strategy, the embryonic development takes place within the body of the mother which provides gas exchange and nutrients for embryos. Here we present the results of histological and EM analyses of the female reproductive system of the viviparous earwig, Arixenia esau, focusing on all the modifications related to the viviparity. We show that in the studied species the embryonic development consists of two "physiological phases" that take place in two clearly disparate compartments, i.e. the terminal ovarian follicle and the uterus. In both compartments the embryos are associated with synthetically active epithelial cells. We suggest that these cells are involved in the nourishment of the embryo. Our results indicate that viviparity in arixeniids is more complex than previously considered. We propose the new term "pseudoplacento-uterotrophic viviparity" for this unique two-phase reproductive strategy.

Hepatitis G virus/GBV-C in serum, peripheral blood mononuclear cells and bone marrow in patients with hematological malignancies.

BACKGROUND: HGV/GBV-C is highly prevalent in the general population but its significance remains unclear. It is known that HGV/GBV-C is not primary hepatotropic and its replication was reported in PBMC, bone marrow and other tissues. To investigate a possible role of HGV/GBV-C 115 consecutive patients with hematological malignancies were analyzed for virus RNA presence and quasispecies composition in three compartments: serum, PBMC and bone marrow. METHODS: RT-PCR was used to amplify 5'UTR HGV/GBV-C in serum, PBMC and bone marrow. Viral sequences obtained from three compartments were subjected for comparative molecular analysis performed by single strand conformational polymorphism (SSCP) and pyrosequencing. RESULTS: HGV/GBV-C RNA was detected in 23 out of 115 (20.0%) patients, most often in bone marrow (18 patients), followed by PBMC (11 patients) and serum (10 patients). Differences in SSCP bands distribution corresponding to different viral variants and confirmed by direct sequencing were observed in three patients. CONCLUSION: HGV/GBV-C infection is frequent in patients with hematological malignancies. Common detection of HGV/GBV-C in bone marrow supports the hypothesis that it is a major replication site of this virus.

Differing strategies of patterning of follicular cells in higher and lower brachycerans (Diptera: Brachycera).

In all higher dipterans (Brachycera), including the fruitfly, Drosophila melanogaster, each egg chamber (ovarian follicle) consists of a group (clone) of germ cells (one oocyte and 15 accompanying nurse cells) that is surrounded by a layer of somatic mesodermal follicular cells (FCs). As oogenesis progresses the initially uniform FCs diversify into several morphologically and functionally distinct subpopulations. In D. melanogaster some of these subpopulations, e.g., border, centripetal, and dorsolateral cells, undertake coordinated migration or rearrangement over the surface of the germ cells. During the final stages of oogenesis these subpopulations participate in the formation of a complex, regionally specialized eggshell. In representatives of lower brachycerans (Orthorrhapha), only FCs that undertake active, directed migration are the border cells. These cells originate at the anterior pole of the ovarian follicle and migrate between the nurse cells to the anterior pole of the oocyte. Reduced motility of FCs in lower brachycerans results in the absence of certain FC subpopulations in their egg chambers and subsequent simplicity of their eggshells. We found that the lack of some FC subpopulations coincided with the appearance of lamellipodium-like protrusions of the oocyte. These protrusions penetrated between the apposing membranes of nurse and FCs and partially enveloped the nurse cell compartment. Analysis of whole-mount preparations stained with rhodamine-conjugated phalloidin revealed that the protrusions contained microfilaments and that their tips were equipped with actin-rich filopodium-like processes. We also found that in some lower brachycerans (representatives of the family Rhagionidae), the FCs located at the posterior pole of the oocyte, became enlarged and morphologically similar to the anterior border cells. These findings indicate that in higher dipterans the processes leading to the formation of a functional egg are variable and often markedly different from those in the model organism, D. melanogaster.