The effects of recombinant interleukin -3 (IL-3) on the myelosuppression induced by irradiation and cyclophosphamide (CY) were observed in mice. The experimental results were as follows: (1) Intraperitoneal (i.p.) or subcutaneous (s.c.) injections of rh IL-3 daily for 5 days immediately after irradiation could alleviate the radiation-induced hematopoietic injuries. The yields and the numbers of CFU-E, BFU-E, CFU-Mix and CFU-GM in femural marrow in mice on the 9th day post exposure to 7 Gy gamma-rays were much higher than those in control animals. Meanwhile, rh IL-3 showed a weak influence on the numbers of bone marrow nucleated cells (BMC) and endogeneous CFU-S. (2) Subcutaneous administration of rh IL-3 for 5 consecutive days brought a favour of CY treated mice to elevate the yields of hematopoietic progenitor cells. (3) The effects of rh IL-3 on myelosuppression induced by radiation or CY were closely related to the route of administration, administration schedules, dosage of this cytokine and the disease state as well. So it seemed important to research still further an appropriate, optimal and flexible guide for clinical use. (4) In vitro rh IL-3 had no effect on the growth of murine BMC and CFU-GM. In comparison with control, after coculture with rmIL-3 (recombinant mouse interleukin (3)) BMCs of normal mice and mice irradiated with 2 Gy gamma-rays proliferated more rapidly and the yield of CFU-GM in them were higher. (5) The mechanism of the effects of rh IL-3 in marrow hypoplastic mice might be related to the indirect promoting influences on the proliferation and/or differentiation of the radiation damaged hematopoietic progenitor cells and/or hematopoietic stem cells.

The effect of ciliary neurotrophic factor (CNTF) on gentamycin induced deafness was observed by Preyer's reflex, auditory brainstem evoked potential, bioelectric response of the cochlea and histomorphological examination of surface preparation of cochlea. It was found that CNTF was capable of reducing ototoxicity of gentamicin in guinea pigs, thus protecting hair cells of cochlea and auditory nerves.

[corrected] To elucidate the role of immunodysfunction in the pathogenesis of severe aplastic anemia (SAA).

To investigate the effect of histamine type II (H2) receptors on the inhibition of hematopoiesis by CD8+ cells from aplastic anemia (AA) patients.

To investigate the pharmaceutical effect of Spatholobus Suberectus Composita (SEC) on bone marrow hematopoiesis and microenvironment of aplastic anemia.

In this study, plasma clot and methycellulose semi-solid and liquid culture technigues were employed to observe the in vitro growth characteristics of proliferation and differetiation of 4-5 months fetal liver and adult bone marrow CFU-MKs. Fetal liver MK colonies in plasma clot in the presence of rIL3 or Meg-CSA were larger (usually with > 50 cells/colony) than that of the adult (usually with < 50 cells/colony). The number (36.40 +/- 16.60/1 x 10(5) cells) of fetal liver Mk colonies in presence of 2 ng/ml rIL3 was larger than that (10.05 +/- 2.81/1 x 10(5) cells) of human adult bone marrow MK colonies (P < 0.01). In contrast, the major DNA ploidy classes of megakaryocytes derived from fetal liver CFU-MK were those of 2N and 4N cells and the major DNA ploidy classes of megakaryocytes derived from human adult bone marrow were those of 8N and 4N cells. The mean (5.45 +/- 0.86) of the ploidy of the former was lower than that (10.13 +/- 1.30) of the latter (P < 0.01). The same results were obtained with the presence of 10% Meg-CSA. These present results indicated that CFU-MK in fetal liver has a high ability of proliferation and low capacity of differentiation (polyploidization). Interestingly, the number of fetal liver MK colonies increased in the range of rIL3 concentration from 0.5 ng/ml to 2 ng/ml. At higher rIL3 concentration (2-8 ng/ml), the colony growth showed a steady decrease from the maximum value instead of an increase. However, in the same range of rIL3 concentration, the numbers of adult bone marrow MK colonies numbers and fetal liver CFU-GM colonies increased steadily and finally reached to a plateau. Furthermore, both fetal liver and adult bone marrow MK colonies showed dose-dependent response in the range of Meg-CSA concentration from 5% to 25%. In addition, there was no difference on DNA ploidy distributions of megakaryocytes derived from fetal liver CFU-MK between rIL3 and Meg-CSA as growth factors. Moreover, the DNA ploidy distribution of fetal liver derived megakaryocytes stimulited by rIL3 could not be changed by addition of rIL6 (100 u/ml). In summary, the above data suggest that CFU-MK in the fetal liver undergoes some intrinsic cellular modification in order to suit the need of ontogensis.

To elucidate the expanding response of cord blood CD34+ cells to human hematopoietic growth factors (HGFs) and evaluate their grafting efficiency.

To explore the cytomorphological and cytochemitry features of CD34+ hematopoietic cells.

To investigate the regulating effect of IL-12 on hemopoiesis.

To show the coexpression levels of P 53, c-myc or bcl-2, three key apoptosis-regulating proteins on CD34+HC and bone marrow mononuclear cells (BMMNC).

To explore the transfer and expression of recombinant human stem cell factor (rh-SCF) gene in human hematopoietic stem/progenitor cell.

The total RNA of HepG2 cell was extracted as the template by ultrocentrifuge method. The full length cDNA (0.8 kb) encoding the human stem cell factor (hSCF) was amplified by RT-PCR method. The cDNA encoding mature hSCF (0.5 kb) was sequenced and was recombined into the expression vector (PBV-220). The expression level of rhSCF in E. coli DH5 alpha was about 15% of the total protein.

C1027, composed of an enediyne chromophore and an apoprotein of 110 amino acid residues, is a new highly potent macromolecular antitumor antibiotic. In order to prepare monoclonal antibodies (McAb) against C1027 by hybridoma technique, natural C1027 was inactivated by ultraviolet and coupled with human serum albumin, then immunized BALB/c mice. After cell fusion and screening by ELISA, a hybridoma secreting anti-C1027 McAb designated as F9 was obtained. McAb F9 specifically reacted with C1027 as determined by immunoblot assay. No obvious difference was observed between the reactivity of McAb F9 to natural C1027 and that of McAb F9 to ultraviolet inactivated C1027. This result indicates that the ultraviolet-sensitive chromophore of C1027 may not participate in the epitope for McAb F9. The isotype of F9 is IgG1 and its affinity constant was found to be (2.2 +/- 0.47) x 10(7) L.mol-1 according to Beatty's method. By clonogenic assay, McAb F9 neither affected the cytotoxicity of C1027 to KB cells nor blocked the activity of the chromophore. McAb F9 also specifically reacted with the recombinant truncated C1027 apoprotein in which 16 amino acid residues at the C terminus were deleted. This study suggests that F9 is a valuable McAb for the research of C1027 apoprotein engineering, C1027 related immunoconjugates and screening of new macromolecular antitumor substances with homology to the protein part of C1027.

The sensitivity to photosensitization mediated by the hematoporphyrin photosensitizer PSD-007 of acute promyelocytic leukemic cell line (HL-60) was compared with normal human hemopoietic progenitor cells. The results showed that the leukemic cells were more sensitive. After being treated with 10 micrograms.ml-1 PSD-007 followed by 2J.cm-2 copper vapor laser light irradiation, the clonogenic leukemic (HL-60) cells were reduced 98%, but the survival rate of human granulocyte-macrophage colony-forming units (CFU-GM) was 40 +/- 8%. Mixing of normal human marrow cells with leukemic (HL-60) cells (ratio 100:1) did not interfere with elimination of tumor cells. The ultrastructure changes of HL-60 cells treated by laser photoradiation was observed under the trans-electronic microscope. The mitochondria, endoplasmic reticulum and cell membrane were involved. This means that the cell biomembrane is the main target to be attacked. It is considered that PSD-007-laser photoradiation therapy is efficient for killing leukemic cells.

The effects of media buffered with HEPES of different concentrations and different incubation periods on the proliferation and differentiation of CFU-GM were studied under the standard condition for culture. The results exhibited that when the incubation period prolonged, the pH values of culture system raised and the colony formation of CFU-GM decreased; the culture system added with proper HEPES buffer solution could maintain and increase the colony formation of CFU-GM, especially the culture system in which the pH value was not adjusted before the experiment. We also found that media buffered with HEPES could change the ratio of different colony types. This showed that media buffered with HEPES could not only stimulate the proliferation of CFU-GM, but also affect its differentiation. The results suggest that media buffered with HEPES are better than those without HEPES for the culture of hematopoietic progenitor cells, and the optimum concentration is 10-20 mmol.L-1.

To assess the efficacy of "method of activating blood circulation and removing stasis (ABCRS)" in treating polycythemia vera.

We constructed plasmids pSVLD(+) and pSVLD(-) containing human D-form Leukemia Inhibitory Factor (LIF) cDNA sequence in sense or antisense orientation, transfected them into cells of an embryonic stem cell line ES-5, and isolated 248 pSVLD(+)-transfected and 93 pSVLD(-)-transfected G 418-resistant clones. By stepwise reducing LIF concentration in the medium, we obtained 3 pSVLD(+)-transfected clones (A 1-3) that could grow in 15% BRL-CM, including ESL(+)A 2 that could grow without LIF: we also obtained 13 pSVLD(-)-transfected clones (B 1-13) which would differentiate in 60% BRL-CM, including ESL(-)B 3 and B 5 that could not be passaged without LIF. ESL(+)A 2 and ESL(-)B 5 cells had the relatively stronger LIF mRNA or antisense LIF RNA expression, and LIF overexpression in ESL(+)A 2 cells was shown by biological assay for ES cell differentiation inhibition. ESL(+)A 2 cells could be continuously passaged for at least 13 passages without addition of exogenous LIF, retained undifferentiated morphology as well as a high growth rate, and resembled ES-5 cells in terms of stem cell characteristics and pluripotent properties, as analyzed for alkaline phosphatase activity and with staining the paraffin sections of tumor formed by inoculating ESL(+) A 2 cells into mouse. On the contrary, ESL(-) cells should be cultured in higher concentration of LIF than ES-5 cells, otherwise, would undertake extensive differentiation. By hanging drop culture for 3 days in the presence of 10(-6) mol/L RA then observing the differentiation of the formed embryonic bodies (EBs), we found that ESL(+) A 2 and ES-5 cells underwent similar morphologically differentiation, with round and epitheliallike cells occurring around the EBs; while ESL(-) B 5 cells, despite initial differentiation to round cells, differentiate into fibroblast-like and spindle shaped cells. The above results indicate that LIF overexpression in ESL(+) A 2 cells is able to completely free ES cells from the dependence on LIF-conditioned medium, and endogenous LIF gene expression, although is very low, may be indispensable for inhibiting the differentiation in vitro of ES cells; LIF overexpression might not obviously change the differentiation way of ES-5 cells, however, blocking endogenous LIF expression gives rise to the increased sensitivity of ES-5 cells to differentiate, with an altered differentiation pattern. The establishment of ESL(+) and ESL (-) cell lines provides models for further study of the growth and differentiation of ES-5 cells.

The Shengxue Syrup (SXS) composed of Chinese medicinal herbs for invigorating the function of Spleen and resplenishing the Kidney was used in treating 115 chronic aplastic anemia patients. The SXS group consisted of 67 patients. After the median 20 months treatment the total effective rate (TER) was 97.0%. The other group was SXS + testosterone group consisted of 48 patients. After median 17.5 month treatment the TER was 95.8%. The difference of TER between two groups was insignificant. The result of animal experiment showed that SXS could markedly enhance the hematopoietic stem cells as well as progenitor cells proliferation of bone marrow in mice.

In order to study the multidifferentiation of medullary carcinoma of the thyroid gland (MCT), 24 cases of MCT were examined for the presence of immunoreactive calcitonin (CT), thyroglobulin (Tg), chromogranin A (CgA), somatostatin (SS), serotonin (5-HT), S-100 protein (S-100), neuron-specific enolase (NSE), vasoactive intestinal polypeptide (VIP), adrenocorticotrophin (ACTH) and neurofilament protein (NF) by using immunohistochemical ABC methods. Results showed that CT-immunoreactive cells were present in all tumors. Tg was present in three tumors. 23 cases contained CgA-immunoreactive cells. 14 tumors contained 5-HT-immunoreactive cells, 10 cases were immunoreactive to NSE and SS. 4 tumors contained VIP-immunoreactive cells and only one cases was positive for S-100. The demonstration of immunoreactivity for multiple antigens in 24 cases suggests that the origin of medullary thyroid carcinoma may originate from neuroectoderm cells potentially capable of producing numerous hormone substances. In addition, as the neoplastic cells in 12% of the tumors containing hormone substances as well as thyroglobulin, it is suggested that follicular epithelial differentiation and mixed medullary thyroid carcinoma may be more common than previously suspected. Recent studies indicate that mixed carcinoma of the thyroid may be derived from common stem cells in posterior branchia capable of differentiating into both follicular and parafollicular tumor cells.

In this study, 16 eligible patients with intermediate and high-grade non-Hodgkin's lymphoma were treated with a new high-dose DHACT regimen supported by rhG-CSF and peripheral blood stem cell (PBSC) rescue. PBSC were mobilized by rhG-CSF or rhGM-CSF. Single leukapheresis was performed and the PBSC were then frozen in liquid nitrogen. CFU-GM clonogenic assay for mononuclear cells and resuscitated progenitor cells done to calculate how many progenitor cells were alive after freezing. The DHACT chemotherapy was composed of carboplatin 600 mg/m2 on d1, Ara-C 1500 mg/m2 on d2, VM-26 100 mg/m2 on d3, 4, and dexamethasone 40 mg/d, on d1-4. Autologous PBSC was reinfused after 24 to 48 hours of chemotherapy. Recombinant human G-CSF at 300 micrograms administered daily on 2 successive days when the absolute neutrophil count was greater than 1 x 10(9)/L. Other supportive care procedures were standard for the unit. The median amount of PBSC reinfused into a patient was 0.9 x 10(8)/kg. The recovery rate of CFU-GM was 78% after cryopresevation. Within 7 to 9 days after high-dose DHACT chemotherapy, the WBC count and the platlet count arrived nadir, and then rose gradually with rhG-CSF injection. The median time for WBC count from nadir to > or = 1 x 10(9)/L was 4 days, and that for platelet count from nadir to > or = 50 x 10(9)/L was 7 days. Nine patients achieved complete remission and 5 patients achieved partial remission. The median follow-up on survival was 9 months. High-dose DHACT regimen supported by rhG-CSF and PBSC rescue is a safe and effective treatment for patients with advanced intermediate and high-grade non-Hodgkin's lymphoma.