Patterns of cellular uptake and effects on cell survival using antimetallothionein oligodeoxyribonucleotide conjugates in vitro

Synthetic oligodeoxyribonucleotides (ODNs) offer the potential for the sequence-specific modulation of viral and cellular gene expression. However, several problems such as efficient delivery into cells, metabolic stability and delivery to specific cellular targets may limit their usefulness. Studies were designed to demonstrate that the covalent conjugation of an 18-mer ODN complementary in sequence to mRNA ODN with various polypeptide ligands, including poly(L-lysine), phosphomannan and asialo-orosomucoid, elicits a pattern of enhanced yet differential uptake into Chang and V79 cells in culture. Viability of cells exposed to conjugated ODNs was measured using a colorimetric assay (MTT). The ODNs covalently linked to poly(L-lysine) reveal an increased efficiency of antisense-directed cell killing from concentrations greater than 3 microM to less than 100 nM. Finally, poly(L-lysine) is also cytotoxic, particularly at extremes of molecular weight. Hence, these studies indicate that synthetic ODNs conjugated to peptides may offer enhanced cellular uptake leading to more efficient antisense activity. However, the cytotoxicity of ODN conjugates may limit their usefulness as research tools or therapeutic agents.     

Erythropoietin mRNA expression in human fetal and neonatal tissue

Based on animal experiments, a switch of the erythropoietin (EPO) production site from the liver in the fetus to the kidneys in the adult has been postulated. To study the switch in humans, we have quantitated EPO mRNA expression in liver, kidney, spleen, and bone marrow of human fetuses and neonates by means of a competitive polymerase chain reaction (PCR). Tissue samples from 66 routine postmortem examinations were obtained. EPO mRNA was expressed in 97% of the tissue specimen derived from the liver (n = 66) and in 93% of those from the kidneys (17 weeks of gestation until 18 months after birth; n = 59). For the first time the EPO gene was found expressed in vivo in human spleen (96% of 64 samples) and in fetal and neonatal bone marrow (81% of 21 samples). EPO mRNA expression in the kidneys increased significantly beyond 30 weeks of gestation (P < .05). Although there was a slight decrease in EPO mRNA content per g liver tissue towards birth, the liver accounted for about 80% of the total body EPO mRNA. The contribution of the spleen and bone marrow were minor compared with liver and kidneys. Our results indicate that in humans the liver is the primary site of EPO gene expression not only in fetal, but also in neonatal life. A significant increase of renal EPO mRNA expression after 30 weeks of gestation might indicate the beginning switch.     

Erythroid colony formation (CFUe) in fetal liver and adult bone marrow and spleen from the mouse

The methyl cellulose modification of the CFUe technique has been applied to 14 day fetal liver and adult bone marrow and spleen from CBA/CA mice. Optimized doses of fetal calf serum, alpha-thioglycerol, erythropoietin and cell suspensions have been obtained from dose response curves in order to standardize the technique. The slopes of the erythropoietin and cell dose response curves indicate a greater sensitivity by fetal liver to the hormone than bone marrow or spleen. The proportion of cells in the DNA synthesis phase of the cell cycle, as measured by the CFUe technique, has been estimated by administering hydroxyurea. Two hours after the drug was injected, 89% of fetal liver cells, 71% of bone marrow cells and 81% of spleen cells were found to be in the S-phase.     

Bone marrow and renal injury associated with haloalkene cysteine conjugates in calves

Almost 40 years ago, it was reported that cattle-feed which had been extracted with hot trichloroethylene and then fed to calves produced renal injury and a fatal aplastic anaemia. The toxic factor was subsequently identified as S-(1,2-dichlorovinyl)-L-cysteine (DCVC). These original findings have been confirmed, a single intravenous dose of DCVC at 4 mg/kg, or 0.4 mg/kg intravenously per day administered for 10 days to calves produced aplastic anaemia, and renal injury after a single dose of 4 mg/kg. The toxicity to calves of a number of other haloalkene cysteine conjugates has been examined to ascertain whether, like DCVC, they produce bone marrow and renal injury. Intravenous administration of the N-acetyl cysteine conjugate of DCVC produced renal but not bone marrow injury at a molar equivalent dose to DCVC, indicating that the calf can deacetylate the mercapturic acid and further that sufficient chemical had reached the kidney to be a substrate for the enzyme cysteine conjugate beta-lyase. However, intravenous administration of the alpha-methyl analogue of DCVC, which cannot undergo metabolism via the enzyme cysteine conjugate beta-lyase, was without toxicity at doses about five-fold higher than DCVC. These latter findings provide strong evidence that metabolism of DCVC via the enzyme beta-lyase is necessary for bone marrow and renal injury to occur. The cysteine conjugates of perchloroethylene and hexachloro-1,3-butadiene(HCBD) when given intravenously to calves at molar equivalent doses to DCVC, or above, did not produce either bone marrow or renal injury. In contrast, intravenous administration of the cysteine conjugate of tetrafluoroethylene (TFEC) produced severe renal tubular injury in calves without affecting the bone marrow. In vitro studies with these haloalkene cysteine conjugates showed, like DCVC, that they were good substrates for calf renal cysteine conjugate beta-lyase and toxic to renal cells as judged by their ability to reduce organic anion and cation transport by slices of calf renal cortex and inhibit the renal enzyme glutathione reductase. Calves were also dosed either orally or intravenously with HCBD to assess its toxicity. HCBD at higher molar equivalent doses than DCVC produced mid-zonal necrosis in the liver, renal tubular necrosis but no bone marrow injury in calves. The key findings emerging from these studies are (1) that none of the other cysteine conjugates, at molar equivalent doses to DCVC and above, produce bone marrow injury in calves, (2) TFEC produced only renal injury, suggesting that sufficient of the other conjugates had not reached the kidney for metabolism by beta-lyase to produce cytotoxicity and (3) that HCBD itself is more toxic than its cysteine or mercapturic acid conjugate, suggesting that pharmaco-kinetics and disposition are important factors in determining the toxicity of these conjugates to calves. Further studies are needed to understand the basis for the selective toxicity of DCVC to the bone marrow of calves.     

Keratinocyte growth factor administered before conditioning ameliorates graft-versus-host disease after allogeneic bone marrow transplantation in mice

Keratinocyte growth factor (KGF) is important in tissue repair and wound healing and its administration can abrogate chemical- and radiation-induced tissue damage in rodents. We investigated KGF as a therapeutic agent for the prevention of graft-versus-host disease (GVHD)-induced tissue damage, morbidity, and mortality in an established murine allogeneic bone marrow transplantation (BMT) model. B10.BR (H2(k)) recipient mice were lethally irradiated and transplanted with C57BL/6 (H2(b)) bone marrow (BM) with spleen cells (BMS) as a source of GVHD-causing T cells. KGF-treated mice (5 mg/kg/d subcutaneously days -6, -5, and -4 pre-BMT) receiving BMS exhibited better survival than those not receiving KGF (P =.0027). Cyclophosphamide (Cy), a common component of total body irradiation (TBI)-containing regimens, was administered to other cohorts of mice at a dose of 120 mg/kg/d intraperitoneally on days -3 and -2 before BMT. KGF-treated mice again exhibited a better survival rate than those not receiving KGF (P =.00086). However, KGF-treated recipients receiving TBI or Cy/TBI BMS were not GVHD-free, as shown by lower body weights compared with BM groups. GVHD target tissues were assessed histologically during a 38-day post-BMT observation period. KGF ameliorated GVHD-induced tissue damage in the liver, skin, and lung (completely in some recipients) and moderately so in the spleen, colon, and ileum, even with Cy conditioning. These studies demonstrate that KGF administration, completed before conditioning, has potential as an anti-GVHD therapeutic agent.     

The hypereosinophilic variant of Ph''-positive chronic myeloleukemia

AIM: To confirm clonal nature of idiopathic hypereosinophilic syndrome (IHES), its relevance to Ph'-positive chronic myeloid leukemia. MATERIALS AND METHODS: 3 cases of idiopathic hypereosinophilic syndrome are reported with morphologic analysis of bone marrow cells and cytogenetic examinations. In one patient the presence of Ph'-chromosome was confirmed at fluorescent in situ hybridization (FISH) and molecular-genetic analysis (bcr/abl). Samples of bone marrow, spleen and liver were examined pathohistologically. RESULTS: The presence of chromosome anomaly t(9;22), i.e. Ph'-chromosome, associated with chronic myeloid leukemia (CML) was identified in all the 3 cases. There was also myeloid hyperplasia in the bone marrow (with primarily mature, eosinophilic granulocytes), spleen and liver, depression of megakaryocyto- and erythropoiesis. 2 patients had similar clinical symptoms which was not typical for CML in chronic phase: fever, elevated ESR, clear-cut anemia and thrombocytopenia. In the absence of hyperleukocytosis, blood and bone marrow eosinophils remained high (42.5, 21.5, 42.5% and 21.4, 7.1, 6.5%, respectively) due to "mature" forms. The number of blasts in the bone marrow was maximum 2.4%. CONCLUSION: The literature and the obtained data suggest closeness of idiopathic hypereosinophilic syndrome and Ph'-positive CML within myeloproliferative diseases.     

Long circulating biodegradable poly(phosphazene) nanoparticles surface modified with poly(phosphazene)-poly(ethylene oxide) copolymer

The biodistribution of biodegradable poly(organo phosphazene) nanoparticles surface modified by adsorption of a novel poly(organo phosphazene)-poly(ethylene oxide) copolymer with a 5000 M(W) PEO chain (PF-PEO[5000]), following intravenous administration in rats and rabbits, is described. The data are compared to the biodistribution of poly(organo phosphazene) and poly(lactide-co-glycolide) nanoparticles coated with a tetrafunctional copolymer of poly(ethylene oxide)-poly(propylene oxide) ethylenediamine, commercially available as Poloxamine 908. This copolymer has a PEO chain of the same size as the poly(organo phosphazene)-PEO derivative used. The results in the rat model reveal that poly(organo phosphazene) nanoparticles with a Poloxamine 908 coating were mainly captured by the liver, although a retardation in clearance from the systemic circulation was seen. In contrast, the poly(organo phosphazene) nanoparticles coated with PF-PEO(5000) showed a prolonged blood circulating profile, with only a small amount of the nanoparticles sequestered by the liver. This indicates the importance of the nature of both the anchoring group and the particle surface on the biological performances of the system. Study of the biodistribution of the PF-PEO(5000)-coated poly(organo phosphazene) nanoparticles in the rabbit model also indicated a prolonged systemic circulation lifetime and reduced liver uptake, whereby a significant amount of the administered nanoparticles was targeted to the bone marrow.     

Lipid-saccharide intermediates in glycoprotein biosynthesis. I. Formation of an oligosaccharide-lipid by thyroid slices and evaluation of its role in protein glycosylation

The synthesis of oligosaccharide-lipids thought to play a role in the attachment of carbohydrate to protein has been studied in incubations of slices from calf kidney, pancreas, thymus, and liver, as well as from hen oviduct. These compounds were characterized after radiolabeling of their saccharide moiety by incubation with [14C]glucose or [14C]mannose and a comparison was made with the oligosaccharide-lipid produced by thyroid slices. Furthermore, the unlabeled glycolipid was prepared from hen oviduct for the purpose of quantitating its sugar constituents. Purification of the oligosaccharide-lipids extracted with chloroform/methanol/water (10/10/3) was achieved by DEAE-cellulose chromatography and their carbohydrate moieties were released by mild acid hydrolysis. On the basis of gel filtration it was determined that the lipid-bound oligosaccharides formed by oviduct, thymus, kidney, and liver had molecular weights comparable to that from thyroid (about 2400). The saccharide moiety of the glycolipid from pancreas was however distinctly smaller in size with a molecular weight of approximately 1800. Analyses of the radiolabeled oligosaccharide-lipids from oviduct, kidney, and thymus indicated that they, like the compound from thyroid slices, but unlike those believed to be formed by cell-free systems from various tissues, contained glucose in addition to mannose and N-acetylglucosamine as their monosaccharide constituents. This compositional data was supported by the finding that the unlabeled oligosaccharide from oviduct consists of 10 mannose, 1 glucose, and 2 N-acetylglucosamine residues. Sodium borohydride reduction of this oviduct saccharide moiety indicated that 1 of the 2 glucosamines was situated in a reducing terminal position. The radiolabeled oligosaccharide from the glycolipid produced by pancreas differed from the others analyzed in that it contained only trace amounts of glucose. Upon treatment with alpha-mannosidase this glucose-deficient pancreatic oligosaccharide was extensively digested (85% of the mannose released). In contrast, the carbohydrate moieties of oviduct, kidney, and thymus, like that of thyroid, underwent a more limited digestion with the alpha-mannosidase (55% or less of the mannose released) suggesting that the presence of glucose may serve to block a more complete degradation of these oligosaccharides by this enzyme.     

Studies of the hemopoietic microenvironments. V. Changes in murine splenic and bone marrow glycosaminoglycans during post irradiation hemopoietic regeneration

Changes in the amount of sulphated and unsulphated glycosaminoglycans in the spleen and the bone marrow of lethally irradiated mice were followed up to 11 days after irradiation. One day after irradiation, a sharp decrease in the amount of glycosaminoglycans was observed. In the absence of hemopoietic cells, the remaining stromal elements of spleen and bone marrow underwent subsequently marked changes in the amount of the sulphated and unsulphated components of the glycosaminoglycans in both organs. Reconstitution of irradiated mice with bone marrow cells affected the pattern of changes in the amount of glycosaminoglycans. Although no linear correlation could be observed between the amount of glycosaminoglycans and the number of hemopoietic cells present in the hemopoietic organs, these results suggest an interaction between hemopoietic cells and stromal cells in the hemopoietic organs with regard to the glycosaminoglycan metabolism.     

Exploratory laparotomy and splenectomy in the diagnosis of Hodgkin''s disease in children

Thirty-three children with histologically proved Hodgkin's disease underwent exploratory laparotomy with splenectomy and biopsy of lumbar lymph nodes, liver, and bone marrow. In 13 patients (39%) the lesion of spleen and/or splenic hilar lymph nodes was diagnosed. The involvement of spleen was mostly found in patients with mixed cellularity type, in the presence of systemic symptoms ("B"), and in the biological stage "b." Involvement of the liver was found in 1 patient, of the hepatic hilar lymph nodes in 3 patients, and of the bone marrow in 4 patients. The lymphographic and histologic data as regards lumbar lymph nodes coincided in 65% of cases. After operation the stage of the disease was changed in 48% patients. The early and late complications were not numerous.     

Toxicity of liposomal 3'-5'-O-dipalmitoyl-5-fluoro-2'-deoxyuridine in mice

Toxicities of 5-fluoro-2'-deoxyuridine (FUdR) and its liposome incorporated dipalmitoyl derivative (FUdR-dipalmitate) to mouse bone marrow, spleen, liver and ileum were compared after treatment for 6 consecutive days. The applied doses of the two formulations, which were shown earlier to have equal antitumor activity in mouse tumor models, were 600 and 2 mumol/kg respectively. When applied in these doses, toxicity to the hemopoietic system, measured as a decreases in progenitor and precursor cells of the erythroid and granuloid/macrophage lineage in bone marrow and spleen, was more severe for FUdR than for liposomal FUdR-dipalmitate. In the liver, mitotic figures, as indicators of cell division, were absent for both drugs while in control livers the number of cells in mitosis was approximately 2%. Toxicity to the ileum was more severe for liposomal FUdR-dipalmitate than for FUdR and was manifested by granulocyte infiltration, the presence of cell debris, loss of columnar epithelial cells and enlarged nuclei with prominent nucleoli in these cells. Thus, by prolonging the retention time of FUdR in vivo, using liposomes as a vehicle and FUdR-dipalmitate as a lipophilic prodrug, the dose-limiting toxicity appears to shift from bone marrow to the gastrointestinal tract.     

Posttranslational modifications of trichomonad tubulins; identification of multiple glutamylation sites

A method of conjugating poly(glutamic acid) poly(phosphorothioate)-cross-linked alkaline phosphatase to maleimide-derivatized immunoglobulin is described. Intramolecular autocatalyzed cross-linking of alkaline phosphatase at 2:1 to 4:1 polymer:enzyme ratios introduced 32-68 thiolates on the surface of the enzyme. Depending on the stoichiometry of polymer to enzyme, the cross-linked alkaline phosphatase retained 75-90% of its native catalytic activity. The cross-linked thiolate-functionalized alkaline phosphatase was conjugated to maleimide-derivatized immunoglobulin. Compared to a control prepared using non-cross-linked alkaline phosphatase, these conjugates were smaller in size and more stable to heat. The enzymatic activity of the cross-linked conjugates after incubation at 45 degrees C and pH 7.5 for 25 days was 35% higher than those of the highest-activity control conjugates. The conjugation process could be controlled by varying the stoichiometries of poly(glutamic acid) poly(phosphorothioate), alkaline phosphatase, and immunoglobulin.     

The sonographic appearance of lesions of the spleen and pancreas in an infant with Langerhans' cell histiocytosis

Langerhans' cell histiocytosis (LCH), the term now used to describe the group of diseases known as histiocytosis X, is a rare disorder of the bone marrow-derived histiocytes that may involve the skin, bone, bone marrow, liver, spleen, lungs, lymph nodes, and rarely the pancreas. Sonographically demonstrable lesions of the spleen and pancreas have not been reported. We present a case of disseminated LCH in a 4-week-old infant to demonstrate the sonographic appearance of splenic and pancreatic lesions that occur with this disease.     

Visceral leishmaniasis in the BALB/c mouse: a comparison of the efficacy of a nonionic surfactant formulation of sodium stibogluconate with those of three proprietary formulations of amphotericin B

In this study, treatment efficacies of a nonionic surfactant vesicle formulation of sodium stibogluconate (SSG-NIV) and of several formulations of amphotericin B were compared in a murine model of visceral leishmaniasis. Treatment with multiple doses of AmBisome, Abelcet, and Amphocil (total dose, 12.5 mg of amphotericin B/kg of body weight) resulted in a significant suppression of parasite burdens in liver (P < 0.0005) and spleen (P < 0.0005) compared with those of controls, with Abelcet having the lowest activity. Only AmBisome and Amphocil gave significant suppression of parasites in bone marrow (compared to control values, P < 0.005). In the acute-infection model, single-dose treatments of SSG-NIV (296 mg of SbV/kg), SSG solution (296 mg of SbV/kg), or AmBisome (8 mg of amphotericin B/kg) were equally effective against liver parasites (compared to control values, P < 0.0005). SSG-NIV and AmBisome treatment also significantly suppressed parasites in bone marrow and spleen (P < 0.005), with SSG-NIV treatment being more suppressive (>98% suppression in all three sites). Free-SSG treatment failed to suppress spleen or bone marrow parasites. Infection status influenced treatment outcome. In the chronic-infection model, the AmBisome single-dose treatment was less effective in all three infection sites and the SSG-NIV single-dose treatment was less effective in the spleen. The results of this study suggest that the antileishmanial efficacy of SSG-NIV compares favorably with those of the novel amphotericin B formulations.     

Effects of interleukin-6 on hematopoiesis in allogeneic and syngeneic bone marrow chimeras

Effects of interleukin-6 (IL-6) on hematopoietic progenitor cells were analyzed in murine bone marrow chimeras. When IL-6 was injected into syngeneic [C3H/He-->C3H/He] bone marrow chimeras from day 1 to day 12, the numbers of highly proliferative potential colony-forming units (CFU-HPP) or colony-forming units mix (CFU-Mix) in spleen cells and bone marrow cells increased on day 14 although there was a marked increase in spleen cells but not in bone marrow cells on day 21. The numbers of CFU-HPP increased in spleen cells from allogeneic [BALB/c-->C3H/He] bone marrow chimeras injected with IL-6 on days 14 and 21. In syngeneic bone marrow chimeras, the numbers of colony-forming units granulocyte/macrophage (CFU-GM) and burst colony-forming units (BFU-E) increased similarly to those of CFU-HPP and CFU-Mix on day 14. On day 21, these were mainly increased in spleen cells. In allogeneic bone marrow chimeras, IL-6 decreased the numbers of CFU-GM and BFU-Mix dose-dependently on day 14. Only 10 micrograms of IL-6 increased the numbers of CFU-GM and BFU-E on day 21. In our previous work, we showed that platelet counts increased on day 14 in syngeneic bone marrow chimeras injected with IL-6, whereas platelet and leukocyte counts increased on days 14 and 24 in allogeneic bone marrow chimeras injected with IL-6, correlating inversely with the numbers of hematopoietic progenitor cells. Overall, primitive hematopoietic progenitors (i.e., CFU-HPP and CFU-Mix) existed primarily in spleen cells of allogeneic bone marrow chimeras on day 14, whereas those in spleen cells of syngeneic bone marrow chimeras were found on day 21. These findings indicate that the effect of IL-6 on hematopoiesis in allogeneic bone marrow chimeras is completely different from that in syngeneic bone marrow chimeras, probably via graft-versus-host reaction (GVHR) but not GVH disease (GVHD).     

Biodegradation and biocompatibility of a guided tissue regeneration barrier membrane formed from a liquid polymer material

Biodegradable barrier films were made by coagulating a solution of poly(DL-lactide) in N-methyl-2-pyrrolidone on porous polyethylene pads wetted with saline solution. The semisolid films were cut into 10 x 10 mm barriers and implanted subcutaneously in rabbits. At monthly intervals, the polymer implant sites were compared histologically to those implanted with USP negative control plastic. The polymer films were retrieved from the surrounding tissue, dried, weighed, and the changes in molecular weight determined using gel permeation chromatography. The molecular weight of the polymer decreased at a relatively constant rate over 5 months; however, no significant mass loss occurred until 5 months postimplantation. Also, no distinct histological differences were noted between the polymer barrier and the control plastic sites until 6 months when histiocytes and multinucleated giant cells showed a modest increase around fragmented polymer films. Similar barrier films also were fitted over naturally occurring buccal dehiscence defects in beagle dogs and the tissue sites compared histologically at 6 months to sham-operated control sites. New bone and dense connective tissues closely approximated segments of the remaining polymer and demonstrated the biocompatibility of the biodegradable films. Histomorphometric analyses of treated sites compared to sham controls showed that the polymer barrier is effective in promoting bone and cementum regeneration in periodontal defects in dogs.     

Abnormal development and differentiation of macrophages and dendritic cells in viable motheaten mutant mice deficient in haematopoietic cell phosphatase

In mice homozygous for the 'viable motheaten' (mev) mutation, numbers of macrophage progenitor cells, particularly monocytes, were markedly increased in the bone marrow and spleen. Increased mobilization of these precursor cells to peripheral tissues and their differentiation to macrophages were evidenced by striking increases in macrophage numbers. Immunohistochemical double staining of tissue sections and flow cytometry analyses of single cell suspensions from these mice demonstrated CD5 (Ly-1)-positive macrophages in the peritoneal cavity, spleen and other tissues. Ly-1-positive macrophage precursor cells were demonstrated in the peritoneal cavity of the mev mice and developed in the omental milky spots. The development of marginal metallophilic and marginal zone macrophages was poor in the splenic white pulp and related macrophage populations were absent in the other lymphoid tissues. The numbers of epidermal Langerhans cells in the skin and T cell-associated dendritic cells in the thymic medulla, lymph nodes, and the other peripheral lymphoid tissues were decreased. However, increased numbers of dendritic cells accumulated in the lungs, liver, and kidneys. These abnormalities in development and differentiation of macrophages and dendritic cells may be ascribed to the deficiency in haematopoietic cell SHP-1 tyrosine phosphatase or may be a secondary consequence of abnormal microenvironments, (either constitutive or in response to inflammatory stimuli) in the haematopoietic and lymphopoietic organs and tissues of these mice.     

Hematopoiesis-promoting activity of rat liver biliary epithelial cells: involvement of a cell surface molecule, liver-regulating protein

Stromal cell lines from bone marrow and other blood-forming organs including fetal liver have been found to support hematopoiesis. In this paper, we demonstrate that rat liver biliary epithelial cells (RLEC), most likely originating from primitive bile ductules, are able to support long-term hematopoietic cell growth as well as burst-forming unit-erythroid (BFU-E) and colony-forming unit-granulocyte/macrophage (CFU-GM) production. RLEC have previously been shown to express a cell surface molecule named liver-regulating protein (LRP), which is involved in the long-term maintenance of hepatocyte functions in a coculture system. In addition, LRP-like molecules have been found in spleen, thymus, lymph nodes, and peripheral blood cells. In the present study, we found that hematopoietic cells and several stromal cell types from bone marrow were LRP-positive, and immunoprecipitation revealed polypeptides similar to those found in RLEC. We then investigated the biological role of LRP on hematopoiesis using short-term RLEC and bone marrow stromal cell culture systems. Addition of specific anti-LRP antibody to both systems reduced hematopoietic cell proliferation and committed progenitor production, whereas it did not directly affect the clonal proliferation and maturation of these progenitors in methylcellulose assays. Moreover, using diffusible chamber cultures that suppress direct contacts with hematopoietic cells, we observed low cell growth and no effect of monoclonal antibody (mAb) L8 treatment. All these results strongly argue for a cell proximity signal mediated by RLEC and bone marrow stromal cells and for the involvement of LRP-like molecules in this signal in liver and bone marrow hematopoietic function.     

In vitro release of 5-fluorouracil with cyclic core dendritic polymer

This paper describes the first synthesis of a series of dendritic polymers with a core of 1,4,7,10-tetraazacyclododecane. This core was allowed to react with methyl acrylate through a Michael addition and was then amidated with ethylenediamine. Repeating the two steps led to controlled molecular weight increasing and branching on the molecular level and produced four direction poly(amide-amine) dendrimers. We successfully synthesized dendrimers from generation 0. 5 to generation 5.5. Each generation was analyzed by Fourier- transform infrared (FT-IR) spectroscopy, 1H NMR and elemental analysis. Titrimetry was also used to determine the number of -NH2 of each full generation (2.0, 3.0, 4.0, 5.0). SEC (size exclusion chromatography) was performed to test the purity of G-3.0, G-4.0 and G-5.0. Parts of the outer layer -NH2 groups of the dendrimers generation 4 and generation 5 were acylated by acetic anhydride. The solubility in water of the dendrimer was thus greatly enhanced. The acetylated dendrimers were then reacted with 1-bromoacetyl-5-fluorouracil to form dendrimer-5FU conjugates. Hydrolysis of the conjugates in a phosphate buffer solution (pH 7.4) at 37 degreesC will release free 5FU. Different generation of dendrimer-5FU conjugates exert marking influence on the amount of 5FU released. The dendritic polymer seems to be a promising carrier for the controlled release of antitumor drugs.