Hexafluoropropylene oxide trimer acid, a perfluorooctanoic acid alternative, induces cardiovascular toxicity in zebrafish embryos.

As an increasingly used alternative to perfluorooctanoic acid (PFOA), hexafluoropropylene oxide trimer acid (HFPO-TA) has been widely detected in global water environments. However, little is known regarding its toxic effects on cardiovascular development. Here, zebrafish embryos were treated with egg water containing 0, 60, 120, or 240 mg/L HFPO-TA. Results showed that HFPO-TA treatment led to a significant reduction in both larval survival percentage and heart rate. Furthermore, HFPO-TA exposure caused severe pericardial edema and elongation of the sinus venous to bulbus arteriosus distance (SV-BA) in Tg (myl7: GFP) transgenic larvae, disrupting the expression of genes involved in heart development and thus causing abnormal heart looping. Obvious sprouting angiogenesis was observed in the 120 and 240 mg/L exposed Tg (fli: GFP) transgenic larvae. HFPO-TA treatment also impacted the mRNA levels of genes involved in the vascular endothelial growth factor (VEGF) pathway and embryonic vascular development. HFPO-TA exposure significantly decreased erythrocyte number in Tg (gata1: DsRed) transgenic embryos and influenced gene expression associated with the heme metabolism pathway. HFPO-TA also induced oxidative stress and altered the transcriptional levels of genes related to cell cycle and apoptosis, inhibiting cell proliferation while promoting apoptosis. Therefore, HFPO-TA exposure may induce abnormal development of the cardiovascular and hematopoietic systems in zebrafish embryos, suggesting it may not be a suitable or safe alternative for PFOA.

Experimental Study of the Efficacy of Sodium Deoxyribonucleate Used in Combination with Co-Transplantation of Mesenchymal and Hematopoietic Stem Cells after Exposure to γ-Radiation.

We studied the possibility of using sodium deoxyribonucleate (Derinat) for improving the efficiency of co-transplantation of mesenchymal (MSC) and hematopoietic stem cells (HSC) to female F1(CBA×C57BL/6) mice with bone marrow aplasia caused by exposure to γ-radiation. It was found that immunomodulator Derinat enhanced the effect of co-transplantation, in particular, triple post-irradiation administration of Derinat accelerated hematopoiesis recovery judging from the parameters of peripheral blood, total cellularity of the bone marrow and spleen, and animal survival. Single or double administration of Derinat prior to irradiation was ineffective. The optimal result was obtained when the following scheme was applied: MSC→HSC with an interval of 48 h starting during the first hours after irradiation and triple administration of Derinat (in 10-15 min, 3 and 7 days after irradiation) in a dose of 3 mg/mouse.

Splenic extramedullary hematopoiesis in dogs is frequently detected on multiphase multidetector-row CT as hypervascular nodules.

Extramedullary hematopoiesis (EMH) is the formation and development of blood cells outside the bone marrow, and in dogs it frequently occurs in the spleen. Although splenic EMH is a relatively common condition, data regarding its appearance in veterinary medicine are lacking. Our aim was to describe different multidetector computed tomographic (MDCT) features of splenic EMH in dogs. In this descriptive retrospective study, dogs with cytological diagnosis of splenic EMH and three-phase MDCT study of the abdomen were included. Multi-detector CT findings recorded were splenomegaly, appearance of the parenchyma, and mean attenuation of the spleen and lesions. Out of 89 dogs included, 55 (62%) presented multifocal nodular aspect, 14 (16%) mass, 12 (13%) diffuse heterogeneous parenchyma, and eight (9%) normal spleen. Most lesions were hyperattenuating to the parenchyma in the arterial (57/89, 64%) and portal (59/89, 66%) phases; whereas in the interstitial phase only 40 of 89 (45%) were hyperattenuating. The mean attenuations of the lesions were higher compared to the values of the adjacent spleen, and the difference of the mean attenuation between the hyperattenuating lesions and the parenchyma was significantly higher in arterial and portal phases than in interstitial phase (P < .0001). The most frequent MDCT aspect of splenic extramedullary hematopoiesis consists of multiple nodules hyperattenuating to the normal spleen, best visualized in the arterial and portal phases.

Effects of abnormal temperature and starvation on the internal defense system of the schistosome-transmitting snail Biomphalaria glabrata.

Climate change may affect the internal defense system (IDS) of freshwater snails, and as a result their capacity to transmit disease. We examined effects of short-term exposure to supra- and sub-optimal temperatures or starvation on 3 parameters of the IDS of the schistosome-resistant Salvador strain of Biomphalaria glabrata - hemocyte concentrations, cell division in the amebocyte-producing organ (APO), and resistance to infection with Schistosoma mansoni. Adult snails were exposed to 1 of 3 temperatures, 20°C, 27°C (controls), or 33°C, for 1 or 2weeks, with food. A fourth group was maintained at 27°C, but without food. Compared to the controls, starved snails had significantly higher hemocyte counts at both 1 and 2weeks, although mitotic activity in the APO was significantly lower at both time periods. Exposure to 20°C or 33°C for 1 or 2weeks did not affect hemocyte numbers. However, APO mitotic activity in snails exposed to 20°C was significantly higher at both 1 and 2weeks, whereas mitotic activity in snails exposed to 33°C was significantly lower at 1week but normal at 2weeks. None of the treatments altered the resistance phenotype of Salvador snails. In a follow-up experiment, exposure to 33°C for 4-5h, a treatment previously reported to both induce expression of heat shock proteins (Hsps) and abrogate resistance to infection, caused immediate upregulation of Hsp 70 and Hsp 90 expression, but did not alter resistance, and Hsp expression levels returned to baseline after 2weeks at 33°C. Results of this study indicate that abnormal environmental conditions can have both stimulatory and inhibitory effects on the IDS in adult B. glabrata, and that some degree of acclimation to abnormal temperatures may occur.

Stable Lentiviral Vector Transfer into Mesenchymal Stem Cells In Vivo.

Green fluorescent protein (eGFP) gene was transferred into mouse mesenchymal stem cells in vivo using a lentiviral vector. In 2 months after injection of the lentivirus into the cavity of the femoral bone, up to 30% fibroblast CFU in the bone marrow of infected mice contained the alien gene. The transferred gene was found in more than 50% of adherent layers of longterm bone marrow cultures formed by mesenchymal stem cells from the infected mice bone marrow; 4% fibroblast CFU obtained from these layers were labeled. Ectopic hemopoiesis foci developed after transplantation of the bone marrow from infected mice under the renal capsule of syngeneic recipients contained bone tissue labeled with the alien gene in 57% cases and labeled fibroblast CFU in 11%. The data confirm the possibility of gene transfer with the lentiviral vectors into the mesenchymal stem cells in vivo.

Thrombosis in Myeloid Malignancies: From CHIP to AML.

The development of myeloid malignancies is a multi-step process starting from pre-malignant stages. Large-scale studies on clonal hematopoiesis of indeterminate potential (CHIP) identified this condition as a risk factor for developing hematologic malignancies, in particular myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML). In parallel, CHIP was found to confer an enhanced thrombotic risk, in particular for cardiovascular diseases. In a similar fashion, in recent years, alongside their life-threatening features, increasing attention has been drawn toward thrombotic complications in myeloid malignancies. Thus, the purpose of this review is to gather a growing body of evidence on incidence, pathogenesis and clinical impact of thrombosis in myeloid malignancies at every step of malignant progression, from CHIP to AML.

Acute arrest of hemopoiesis presenting as severe aplastic anemia: a retrospective analysis.

OBJECTIVE: The clinical manifestations of acute arrest of hemopoiesis (AAH) are very similar with severe aplastic anemia (SAA). Currently there are no clear diagnostic criteria to distinguish AAH from SAA. Differentiation of AAH from SAA is challenging in the routine clinical practice. This study aimed to analyze the clinical and laboratory features between AAH and SAA patients. PATIENTS AND METHODS: We performed a retrospective study with cohort of 425 suspected patients who were hospitalized to the First Affiliated Hospital of Zhengzhou University from 1 January 2019 to 31 December 2020. We identified 11 AAH patients and 49 SAA patients to investigate the differentiation diagnostic features. RESULTS: Clinical and laboratory examinations of 11 patients with AAH met the diagnostic criteria of SAA, and hematopoietic recovery occurred within a median time of 12 (4-21) days. The median time for neutrophils to recover above 1 × 109/L and platelet to recover above 50 × 109/L in all patients with AAH was 5 (3-8) days and 8 (1-13) days, respectively. Compared with the control group SAA, the 11 AAH patients were older, with a median age of 53 (21-69) years old, and their first symptom is usually fever. CONCLUSIONS: The spontaneous remission of AAH was rapid in most patients, and relapses were rarely observed. With supportive treatment, the AAH patients would show significant improvement on blood routine about a week, otherwise the patients should be treated as early as possible with the SAA regimen.

Evaluation of bone marrow hemopoiesis and the elemental status of the red bone marrow of chickens under introduction of copper to the organism.

The role of chemical elements in an organism is versatile and multifunctional. However, you should pay attention to the reaction of the organism on the introduction of chemical elements with different biological roles, which is predetermined by the physiological role of organs and body systems. These include the red bone marrow, which primarily responds to endogenous and exogenous factors by its functional significance. Analyzing the myelogram of birds after the various ways of copper NP introduction into the body and the different dosages, we found that, by the end of the experiment, the total numbers of bone marrow cells in all groups were lower than the initial values: in the second group-12.54% lower (p < 0.05), in third-26.32% lower (p < 0.001), for the fourth-14.75% lower (p < 0.05), with exception for the first experimental group where this index was 45.51% higher (р < 0.001). We revealed the following changes in the peripheral blood: the hemoglobin content by the end of the experiment was significantly higher than the initial values: by 18.63% for the first group (p < 0.01); 28.61% higher in the third group (p < 0.001); and 15.76% higher for the fourth (p < 0.01), except the animals of the second group (3.23% lower). The concentration of erythrocytes in all groups was higher than that of the background: by 24.56% (p < 0.001), by 3.37%, by 26.18% (p < 0.001), and by 14.85% (p < 0.01), respectively; the leukocyte concentration in the first group was 39.63% higher (p < 0.001), it remained at the level of the initial values in the other groups. The erythrocyte sedimentation rate in all groups increased by 2.4, 4.0, 2.01, and 1.86 times (p < 0.001), respectively. We revealed that the introduction of copper into an organism in the form of nanopowder both with feed and intramuscularly significantly caused an increase of the content of such elements as arsenic, copper, and silicon and a decrease of calcium, potassium, magnesium, phosphorus, boron, cobalt, iodine, lithium, sodium, zinc, tin, and strontium in the marrowy aspirate. Moreover, compared with the first group (p < 0.01), increasing doses of nanopowders caused a significant rise in the arsenic and tin concentrations and a decline of iodine and strontium. We found that copper nanoparticles ambiguously affect the bone marrow hemopoiesis of poultry; increasing the dose and changing the type of introduction activating the bone marrow hematopoietic function, in particular, granulocyto-, megakaryocyto-, and erythropoiesis.

A comparison of computed tomography with magnetic resonance imaging for the diagnosis of thoracic extramedullary hemopoiesis in patients with leukemia: A non-inferiority retrospective diagnostic study.

At present, MRI is the primary choice of examination for the diagnosis of thoracic extramedullary hemopoiesis. When thoracic extramedullary hemopoiesis presents as posterior mediastinum masses in specific clinical contexts, the diagnosis is not challenging. Other radiological presentations may be more difficult for diagnosis and require biopsy. Needle biopsy is typically preferred for the diagnosis of extramedullary hemopoiesis however, the high vascularization of tissues is one of the complications of this method thus, it is avoided. The aim of the present study was to compare the diagnostic parameters of CT with MRI for the diagnosis of thoracic extramedullary hemopoiesis in patients with leukemia, with an open lung biopsy as a reference standard. Chest CT, chest MRI and open lung biopsy data from a total of 912 patients with leukemia with a sign(s) and symptoms of suspected paravertebral and/or pulmonary extramedullary hemopoiesis were reviewed. In the present study, thoracic extramedullary hemopoiesis was defined as diffusivity of both lung fields being increased compared with the blood pool and no other abnormal focal of lungs being increased compared with the blood pool. The beneficial score was calculated for CT and MRI and plotted for the decision making of irradiation. With respect to open lung biopsy, MRI had a higher sensitivity compared with CT (0.865 vs. 0.809; P<0.0001; q=1691) however, CT had a higher accuracy compared with MRI (0.833 vs. 0.733; P<0.0001; q=3020). The low rate of overdiagnosis was observed for both methods for the detection of thoracic extramedullary hemopoiesis however, the working area for detecting thoracic extramedullary hemopoiesis at least once in images was higher for MRI compared with CT. CT and MRI both have diagnostic importance in the detection of thoracic extramedullary hemopoiesis in patients with leukemia however, chest MRI misdiagnoses the condition while CT can confirm it (level of evidence, 3).

Liposomal Formulation of a Melphalan Lipophilic Prodrug: Studies of Acute Toxicity, Tolerability, and Antitumor Efficacy.

BACKGROUND: Recently we developed a scalable scheme of synthesis of melphalan ester conjugate with 1,2-dioleoyl-sn-glycerol (MlphDG) and a protocol for the fabrication of its lyophilized liposomal formulation. OBJECTIVE: Herein we compared this new convenient in use formulation of MlphDG with parent drug Alkeran® in rats concerning several toxicological parameters and evaluated its antitumor efficacy in the model of breast cancer in mice. METHOD: Liposomes of approximately 100 nm in diameter, consisting of egg phosphatidylcholine, soybean phosphatidylinositol, and MlphDG, or placebo liposomes without the drug were produced by extrusion and lyophilized. Alkeran® or liposomes recovered by the addition of water were injected into the tail vein of animals. Clinical examination of rats consisted of detailed inspection of the behavior, general status, and hematological parameters. Mice with transplanted breast cancer WNT-1 were subjected to multiple treatments with the drugs; tumor growth inhibition was assessed, together with cellular immunity parameters. RESULTS: Liposomes showed approximately two times lower acute toxicity and better tolerability than Alkeran® in terms of behavioral criteria. The toxic effects of liposomes on hemopoiesis were manifested at higher doses than in the case of Alkeran®, proportionally to the difference in LD50 values. The formulation inhibited tumor growth significantly more effectively than Alkeran®, delaying the start of the exponential growth phase and exhibiting no additional toxic effects toward bone marrow. CONCLUSION: Lower toxicity of the liposomal formulation of MlphDG promises improved quality of life for cancer patients in need of treatment with melphalan. Presumably, the list of indications for melphalan therapy could be extended.

Protective effects of squid ink extract towards hemopoietic injuries induced by cyclophosphamine.

To investigate the protective effects of squid ink in chemotherapy, BALB/c mice were used as animal models of injuries induced by cyclophosphamine, a well known chemotherapeutic drug. The mice were randomly divided into five groups with the same number of males and females in each group. At the end of the experiment, animals were sacrificed to investigate organ indexes and antioxidant ability of the spleen, peripheral blood profile and quantities of bone marrow nucleated cells. Results showed that the hemopoietic function of mice was injured by cyclophosphamine, as indicated by decreases of contents of erythrocytes, leukocytes, hemoglobin and bone marrow nucleated cells (P<0.01), while platelets were not affected (P>0.05), as well as modification of organ indexes (P<0.05) and spleen antioxidant ability (P<0.05 or P<0.01), whereas sepia extract markedly increased the levels of erythrocytes, leukocytes, hemoglobin and bone marrow nucleated cells (P<0.01), but not platelets (P>0.05), and reversed the effects of cyclophosphamine on organ indexes and antioxidant ability of spleen (P<0.01 or P<0.05). In addition, squid ink extract did not change marrow hemopoiesis but improved the antioxidant ability of spleen in the animals. The data suggest that squid ink extract can protect the hemopoietic system from chemotherapeutic injury and could be employed to develop cell-protective drugs for use in clinical treatment of tumours.

Five Active Components Compatibility of Astragali Radix and Angelicae Sinensis Radix Protect Hematopoietic Function Against Cyclophosphamide-Induced Injury in Mice and t-BHP-Induced Injury in HSCs.

Although the compatibility of Astragali Radix (AR) and Angelicae Sinensis Radix (ASR) has favorable effect on promoting hematopoiesis in traditional Chinese medicine (TCM), the main active components and pharmacological mechanism are unknown. We investigated the five active components and its mechanisms in vitro and in vivo. Five active components of Astragalus glycosides (AST), Formononetin (FRM), Ferulic acid (FRA), Calycosin (CAL), and Calycosin-7-glucoside (CLG), which could be absorbed in intestinal tract, were detected in this study. The peripheral blood, hematopoietic growth factors (HGFs), and hematopoietic progenitor cells (HPCs) colony were observed to evaluate the effect of these five active components promoting hematopoiesis. Furthermore, hematopoietic stem cell (HSC) proliferation, aging, cycle, and related proteins were detected to explore the mechanism of these five components promoting HSC proliferation. i) The in vivo experiments showed that the combination of the five active components could remarkably increase the number of RBCs, WBCs, PLTs, and content of Hb in peripheral blood and the area of bone marrow hematopoietic tissue, as well as thrombopoietin (TPO), erythropoietin (EPO), granulocyte-macrophage colony stimulating factor (GM-CSF), and colony of CFU-GM, CFU-MK, CFU-E, and BFU-E in serum. Each of these five components promoted the recovery of RBCs and Hb, and increased TPO, CFU-MK, and CFU-E. All components except for AST increased the CFU-GM. FRA increased the number of WBCs, the area of bone marrow hematopoietic tissue, and BFU-E. FRA and AST promoted PLT recovery. FRA and CAL improved the content of GM-CSF. FRA, CAL, and CLG improved the content of EPO. ii) The in vitro experiments showed that FRA, FRM, and AST significantly promoted cell proliferation, reduced the positive rate and G0/G1 cells, and increased G2/M + S cells and the expression of cyclin D1 and CDK4 proteins in aging HSCs. Furthermore, the combination of five components had the best effect. Taken together, the five active components of AST, FRM, FRA, CAL, and CLG were the main pharmacodynamic substances of the AR-ASR compatibility, which promoted hematopoiesis. The combination of them had a synergistic effect. The mechanism of promoting hematopoiesis may be relevant to regulating cyclin-related proteins, promoting cell cycle transformation, and promoting HSC proliferation.

Hematopoietic effect of small molecular fraction of Polygoni multiflori Radix Praeparata in cyclophosphamide-induced anemia mice.

The aim of this study is to investigate the protective effects of a small molecular fraction (SMF) of Polygoni multiflori Radix Praeparata (PMRP) in a cyclophosphamide (CTX) induced anemia mouse model. Small molecular fraction of PMRP was prepared and identified by high-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (HPLC-Q-TOF-MS). In pharmacology, we examined the peripheral hemogram and thymus and spleen index. The content of granulocyte-macrophage colony-stimulating factor (GM-CSF) in serum was mensurated by enzyme-linked immunosorbent assay (ELISA); The level of superoxide dismutase (SOD), catalase (CAT), total antioxidant capacity (T-AOC), and malondialdehyde (MDA) in serum and spleen tissue homogenate were detected, and glutathione peroxidase (GSH-PX) was assayed in spleen. The results show that SMF can significantly accelerate the recovery of peripheral hemogram, increase the activity of antioxidant enzymes and GM-CSF in serum and spleen. SMF also increases the number of spleen cells, improves bone marrow pathology. In conclusion, the SMF of PMRP promoted the recovery of hematopoietic function in a CTX-induced anemia mouse, which can support SMF to be used as an adjunct to chemotherapy to counteract its side effects.

Systems-Based Interactome Analysis for Hematopoiesis Effect of Angelicae sinensis Radix: Regulated Network of Cell Proliferation towards Hemopoiesis.

OBJECTIVE: To explore the molecular-level mechanism on the hematopoiesis effect of Angelicae sinensis Radix (ASR) with systems-based interactome analysis. METHODS: This systems-based interactome analysis was designed to enforce the workflow of "ASR (herb)→compound→target protein→internal protein actions→ending regulated protein for hematopoiesis". This workflow was deployed with restrictions on regulated proteins expresses in bone marrow and anemia disease and futher validated with experiments. RESULTS: The hematopoiesis mechanism of ASR might be accomplished through regulating pathways of cell proliferation towards hemopoiesis with cross-talking agents of spleen tyrosine kinase (SYK), Janus kinase 2 (JAK2), and interleukin-2-inducible T-cell kinase (ITK). The hematopoietic function of ASR was also validated by colony-forming assay performed on mice bone marrow cells. As a result, SYK, JAK2 and ITK were activated. CONCLUSION: This study provides a new approach to systematically study and predict the therapeutic mechanism for ASR based on interactome analysis towards biological process with experimental validations.

Design and Application of Multiplex PCR Seq for the Detection of Somatic Mutations Associated with Myeloid Malignancies.

Targeted sequencing, in which only a selected set of genomic loci are sequenced, enables a much higher coverage of each target than what is obtained using whole genome or exome sequencing. Multiplex PCR offers a simple and affordable technique for specific capture of target regions and can be easily adapted to generate next-generation sequencing (NGS)-ready amplicons. Here we describe a multiplex PCR (MxPCR) approach for capturing 13 leukemia-associated mutation hotspots followed by MiSeq sequencing that enables robust detection of mutations with a variant allele fraction (VAF) as low as 0.8% (0.008) in blood DNA.

Black Scorpionfish (Scorpaena porcus) Hemopoiesis: Analysis by Flow Cytometry and Light Microscopy.

Cell suspensions of head kidney and spleen of black scorpionfish (Scorpaena porcus L.) have been studied using flow cytometry and light microscopy. On the basis of forward scatter (FS) and side scatter (SS) distribution and light microscopy, two main types of cells in the hemopoietic organs were identified: "small cells" (5.1-8.9 µm) and "large cells" (10.8-15.3 µm). Subpopulation of small cells was formed by thrombocytes, lymphocytes, and elements with functionally inactive nuclei. Euchromatin level in their nuclei was low and acidophilic cytoplasm corresponded to negligible nucleic acids content. No proliferative activity was observed using SYBR Green I fluorescence analysis. Morphological characteristics of these cells coincided with colony forming units of mammals. Large cells in head kidney consisted of two subpopulations of cells differing in granularity and DNA content. Proliferating blast cells and differentiating cells of all hemopoietic lines were identified among them. Macrophages and apoptotic cells were also detected in head kidney. In spleen large cells cluster mainly included aged red blood cells with extended lengthwise axis. Blast cells and differentiating elements in spleen were not observed. Anat Rec, 2017. © 2017 Wiley Periodicals, Inc. Anat Rec, 300:1993-1999, 2017. © 2017 Wiley Periodicals, Inc.

Nuclear microRNAs in normal hemopoiesis and cancer.

Since the discovery of microRNAs (miRNAs) in the early 1990s, these small molecules have been increasingly recognized as key players in the regulation of critical biological processes. They have also been implicated in many diverse human diseases. The canonical function of miRNAs is to target the 3' untranslated region (3' UTR) of cytoplasmic messenger RNA to post-transcriptionally regulate mRNA and protein levels. It has now been shown that miRNAs can also bind to the promoter regions of genes or primary miRNA transcripts to regulate gene expression. Such observations have indicated the presence of miRNAs in the nucleus and implied additional non-canonical functions. Nevertheless, the role(s) of nuclear miRNAs in normal hemopoiesis and cancer remains elusive despite a burgeoning literature. Herein, we review current knowledge concerning the abundance and/or functions of nuclear miRNAs during blood cell development and cancer biology. We also discuss ongoing challenges in order to provoke further studies into identifying key roles for nuclear miRNAs in the development of other cell lineages and human cancers.

Microenvironmental Control of Inflammatory Cell Differentiation.

A wide body of information now exists on hemopoietic and proinflammatory cytokine production by structural cells of the microenvironment. Included among these are epithelial cells, endothelial cells and fibroblasts which can, either constitutively or upon stimulation with other cytokines or lipopoly-saccharide, express the genes for, and produce, IL-6, IL-8, G-CSF, GM-CSF, and M-CSF, as well as yet unidentified cytokines with prominent cell differentiation-inducing activities. Inflammatory cells which accumulate at sites of allergic-type reactions include granulocytes such as basophils, eosinophils and mast cells, as well as neutrophils and cells of the monocyte-macrophage lineage. Combinations of cytokines produced by tissue structural cells have been studied with reference to their capacity to induce differentiation, activate and prolong the survival of inflammatory cells. Evidence can be adduced for the differentiation process being intimately connected to phenotype switch and activation of cells, such as eosinophils and mast cells, which themselves can feed back upon this by production of cytokines such as TGF-ß and GM-CSF; the production of T cell-derived cytokines such as IL-3, IL-5 and GM-CSF can be shown to contribute to basophil and eosinophil differentiation and activation. Work from our laboratory will be summarized with reference to the syntax and language of structural cell-derived cytokines in terms of inflammatory cell differentiation pathways, using a variety of in vitro and in vivo detection techniques. Application of these findings to the control of inflammatory reactions as well as wound repair will also be discussed.

Evolution of a cellular immune response in Drosophila: a phenotypic and genomic comparative analysis.

Understanding the genomic basis of evolutionary adaptation requires insight into the molecular basis underlying phenotypic variation. However, even changes in molecular pathways associated with extreme variation, gains and losses of specific phenotypes, remain largely uncharacterized. Here, we investigate the large interspecific differences in the ability to survive infection by parasitoids across 11 Drosophila species and identify genomic changes associated with gains and losses of parasitoid resistance. We show that a cellular immune defense, encapsulation, and the production of a specialized blood cell, lamellocytes, are restricted to a sublineage of Drosophila, but that encapsulation is absent in one species of this sublineage, Drosophila sechellia. Our comparative analyses of hemopoiesis pathway genes and of genes differentially expressed during the encapsulation response revealed that hemopoiesis-associated genes are highly conserved and present in all species independently of their resistance. In contrast, 11 genes that are differentially expressed during the response to parasitoids are novel genes, specific to the Drosophila sublineage capable of lamellocyte-mediated encapsulation. These novel genes, which are predominantly expressed in hemocytes, arose via duplications, whereby five of them also showed signatures of positive selection, as expected if they were recruited for new functions. Three of these novel genes further showed large-scale and presumably loss-of-function sequence changes in D. sechellia, consistent with the loss of resistance in this species. In combination, these convergent lines of evidence suggest that co-option of duplicated genes in existing pathways and subsequent neofunctionalization are likely to have contributed to the evolution of the lamellocyte-mediated encapsulation in Drosophila.

The specifically enhanced cellular immune responses in Pacific oyster (Crassostrea gigas) against secondary challenge with Vibrio splendidus.

The increasing experimental evidences suggest that there are some forms of specific acquired immunity in invertebrates, but the underlying mechanism is not fully understood. In the present study, Pacific oyster (Crassostrea gigas) stimulated primarily by heat-killed Vibrio splendidus displayed stronger immune responses at cellular and molecular levels when they encountered the secondary challenge of live V. splendidus. The total hemocyte counts (THC) increased significantly after the primary stimulation of heat-killed V. splendidus, and it increased even higher (p < 0.01) and reached the peak earlier (at 6 h) after the secondary challenge with live V. splendidus compared with that of the primary stimulation. The number of new generated circulating hemocytes increased dramatically (p < 0.01) at 6 h after the pre-stimulated oysters received the secondary stimulation with live V. splendidus, and the phagocytic rate was also enhanced significantly (p < 0.01) at 12 h after the secondary stimulation. Meanwhile, the enhanced phagocytosis of hemocytes was highly specific for V. splendidus and they could distinguish Vibrio anguillarum, Vibrio coralliilyticus, Yarrowia lipolytica, and Micrococcus luteus efficiently. In addition, the mRNA expression of 12 candidate genes related to phagocytosis and hematopoiesis were also monitored, and the expression levels of CgIntegrin, CgPI3K (phosphatidylinositol 3-kinase), CgRho J, CgMAPKK (mitogen-activated protein kinase kinase), CgRab32, CgNADPH (nicotinamide adenine dinucleotide phosphate) oxidase, CgRunx1 and CgBMP7 (bone morphogenetic protein 7) in the hemocytes of pre-stimulated oysters after the secondary stimulation of V. splendidus were higher (p < 0.01) than that after the primary stimulation, but there was no statistically significant changes for the genes of CgPKC (protein kinase C), CgMyosin, CgActin, and CgGATA 3. These results collectively suggested that the primary stimulation of V. splendidus led to immune priming in oyster with specifically enhanced phagocytosis and rapidly promoted regeneration of circulating hemocytes when the primed oysters encountered the secondary challenge with V. splendidus.