Effect of a neutrophilia-inducing tumor on hemopoietic stem cells in mice

The influence of neutrophilic stimulation on hemopoietic stem cells was studied in mice with tumor-induced neutrophilia. Transfusions of marrow cells from normal and neutrophilic tumor-bearing mice into lethally irradiated normal and tumor-bearing mice were performed. The number and the erythroid:granuloid (E:G) ratio of day 7 colonies in the recipient spleens and bones as well as the size of spleen colonies of recipient animals were determined. The E:G ratio of spleen and bone marrow colonies between normal and tumor-bearing mouse recipients and the number of spleen colonies did not differ significantly in either experiment. However, spleen colonies which developed in tumor-bearing irradiated mice were significantly larger than those which developed in normal recipients in both experiments. These studies indicated that while the line of differentiation taken by hemopoietic stem cells was not affected by the neutrophilic influence of the tumor, the tumor-bearing host environment appeared to enhance proliferation of transfused stem cells and/or their descendants. The stimulators of granulocytopoiesis in this model of neutrophilia appear to act on a population of progenitor cells more mature than the stem cells capable of forming 7-day colonies in the spleen and bone marrow of irradiated recipient mice.     

Components of the hematopoietic compartments in tumor stroma and tumor-bearing mice

Solid tumors are composed of cancerous cells and non-cancerous stroma. A better understanding of the tumor stroma could lead to new therapeutic applications. However, the exact compositions and functions of the tumor stroma are still largely unknown. Here, using a Lewis lung carcinoma implantation mouse model, we examined the hematopoietic compartments in tumor stroma and tumor-bearing mice. Different lineages of differentiated hematopoietic cells existed in tumor stroma with the percentage of myeloid cells increasing and the percentage of lymphoid and erythroid cells decreasing over time. Using bone marrow reconstitution analysis, we showed that the tumor stroma also contained functional hematopoietic stem cells. All hematopoietic cells in the tumor stroma originated from bone marrow. In the bone marrow and peripheral blood of tumor-bearing mice, myeloid populations increased and lymphoid and erythroid populations decreased and numbers of hematopoietic stem cells markedly increased with time. To investigate the function of hematopoietic cells in tumor stroma, we co-implanted various types of hematopoietic cells with cancer cells. We found that total hematopoietic cells in the tumor stroma promoted tumor development. Furthermore, the growth of the primary implanted Lewis lung carcinomas and their metastasis were significantly decreased in mice reconstituted with IGF type I receptor-deficient hematopoietic stem cells, indicating that IGF signaling in the hematopoietic tumor stroma supports tumor outgrowth. These results reveal that hematopoietic cells in the tumor stroma regulate tumor development and that tumor progression significantly alters the host hematopoietic compartment.     

Mobilization of bone marrow stem cells with StemEnhance® improves muscle regeneration in cardiotoxin-induced muscle injury

Bone marrow-derived stem cells have the ability to migrate to sites of tissue damage and participate in tissue regeneration. The number of circulating stem cells has been shown to be a key parameter in this process. Therefore, stimulating the mobilization of bone marrow stem cells may accelerate tissue regeneration in various animal models of injury. In this study we investigated the effect of the bone marrow stem cells mobilizer StemEnhance (SE), a water-soluble extract of the cyanophyta Aphanizomenon flos-aquae (AFA), on hematopoietic recovery after myeloablation as well as recovery from cardiotoxin-induced injury of the anterior tibialis muscle in mice. Control and SE-treated female mice were irradiated, and then transplanted with GFP+ bone marrow stem cells and allowed to recover. Immediately after transplant, animals were gavaged daily with 300 mg/kg of SE in PBS or a PBS control. After hematopoietic recovery (23 days), mice were injected with cardiotoxin in the anterior tibialis muscle. Five weeks later, the anterior tibialis muscles were analyzed for incorporation of GFP+ bone marrow-derived cells using fluorescence imaging. SE significantly enhanced recovery from cardiotoxin-injury. However, StemEnhance did not affect the growth of the animal and did not affect hematopoietic recovery after myeloablation, when compared to control. This study suggests that inducing mobilization of stem cells from the bone marrow is a strategy for muscle regeneration.     

5-Fluorouracil treatment after irradiation impairs recovery of bone marrow functions

Summary In mice, persisting radiation-induced growth retardation of hematopoietic tissue suggested that at least part of the surviving stem cells are genetically injured. Additional mitotic stress some time after the radiation insult might remove injured stem cells, thus improving the overall recovery of the irradiated bone marrow.Mice were treated with 5 Gy whole-body gamma irradiation. Two weeks later half of the animals were injected i.v. with 150 mg/kg 5-fluorouracil (5-FU), the other half remained untreated (5 Gy-controls). 2 or 10 weeks later, femoral cellularity and CFU-S content, proliferation ability of transplanted bone marrow and the compartment ratio (CR; ratio of splenic IUdR incorporation at day 3 and number of CFU-S transfused) were determined.Four weeks after 5 Gy and 2 weeks after 5-FU treatment all parameters showed significant impairment of recovery. 12 weeks after 5 Gy and 10 weeks after 5-FU CFU-S and CR were still reduced compared to the 5 Gy-controls. 5-FU treatment of unirradiated mice did not produce permanent effects on the quality of stem cells or the hematopoietic microenvironment. It is concluded, therefore, that an increased proliferation stimulus does not aid in the removal of injured CFU-S and may even impair recovery of bone marrow functions by increasing the proportion of genetically injured stem cells which continue proliferation.Dedicated to Prof. L.E. Feinendegen on the occasion of his 60th birthday     

Duration of opiate receptor blockade determines tumorigenic response in mice with neuroblastoma: a role for endogenous opioid systems in cancer

The relationship between the pharmacological properties of an opioid antagonist, naltrexone (NTX), and tumor response was studied in mice with transplanted neuroblastoma (NB). Animals receiving 0.1 mg/kg NTX every 6 hr, which blocked morphine-induced analgesia for 24 hr each day, had a 100% tumor incidence, no deviation in time before tumor appearance, and a 17% decrease from control values in total survival time. In contrast, once daily injections of either 0.1 mg/kg NTX or 0.4 mg/kg NTX (the equivalent of 0.1 mg/kg given 4 times daily), which blocked morphine-induced analgesia for less than 10 hr each day, resulted in a tumor incidence of 20% and 60%, respectively, delays in time prior to tumor appearance of 90% and 65%, respectively, and an increased total survival time of 10% and 24%, respectively, for tumor-bearing mice relative to control levels. Inoculation of NB in control animals resulted in 100% tumor appearance within 16 days and a mean survival time of 36 days. These results show that tumorigenic events are dictated by the duration of opiate receptor blockade rather than the dosage of opiate antagonist, and provide compelling evidence that endogenous opioid systems play a crucial role in neuro-oncogenic expression.     

Age-related changes in hematopoietic stem cell populations of a long-lived hybrid mouse

Age-related changes in the number and concentration of pluripotential and unipotential hematopoietic stem cells in the femoral bone marrow and spleen of BC3F₁ mice were investigated. Pluripotential stem cells were assayed by the spleen colony technique, and unipotential stem cells were determined by an agar cloning method and by erythropoietin responsiveness in polycythemic mice. Changes with senescence were observed in the concentration of both uni- and pluripotential stem cells in the bone marrow; the size of the stem cell compartment in the marrow did not change significantly with age. Also, a reduction in the seeding of transplanted spleen colony-forming units into the spleens of aged recipients was demonstrated. The implications of these findings for the kinetics of hematopoietic stem cell proliferation in aged animals are discussed.     

γ-聚谷氨酸-D-半乳糖酯化衍生物-顺铂复合物的制备及其肝靶向性分析

制备一种γ-聚谷氨酸-D-半乳糖酯化衍生物-顺铂复合物［Poly (γ-glutamic acid)D-galactose esterifiable derivative-Cisplatin Complex Compound,γ-D+-DDP］,并考察其体内靶向性。通过生物发酵获得大分子γ-聚谷氨酸［Poly (γ-glutamic acid),γ-PGA］,利用酸降解得到可以作为药物载体的小分子γ-聚谷氨酸;利用凝胶色谱柱检验小分子γ-聚谷氨酸的分子量;利用HPLC检测释放的游离顺铂含量,得到复合物在小鼠体内靶向性分布情况;利用HE组织切片染色,观察脏器受损伤情况及靶向分布。实验结果表明：成功获得γ-聚谷氨酸-D-半乳糖酯化衍生物-顺铂复合物,该复合物载药率达9.4%~10.2%;HPLC结果表明注射复合物后,肝脏中药物显著增加而肾脏中药物分布明显减少,大大减少了肾毒性,肝靶向作用明显。因此,γ-聚谷氨酸-D-半乳糖酯化衍生物-顺铂复合物是一种有效的具有肝靶向性的抗肿瘤药物,具有潜在的临床应用价值;通过生物发酵获得的γ-聚谷氨酸可用于药物载体,赋予药物新的特点。介绍了制备一种γ-聚谷氨酸-D-半乳糖酯化衍生物-顺铂复合物[Poly (γ-glutamic acid)-D-galactose esterifiable derivative -Cisplatin Complex Compound,（γ-D+-DDP）],并考察其体内靶向性。通过生物发酵获得大分子γ-聚谷氨酸[Poly (γ-glutamic acid),γ-PGA],利用酸降解得到可以作为药物载体的小分子γ-聚谷氨酸;利用凝胶色谱柱检验小分子γ-聚谷氨酸的分子量;利用HPLC检测释放的游离顺铂含量,得到复合物在小鼠体内靶向性分布情况;利用HE组织切片染色,观察脏器受损伤情况及靶向分布。实验结果表明：成功获得γ-聚谷氨酸-D-半乳糖酯化衍生物-顺铂复合物,该复合物载药率达9.4% ~10.2%;HPLC结果表明注射复合物后,肝脏中药物显著增加而肾脏中药物分布明显减少,大大减少了肾毒性,肝靶向作用明显。因此,γ-聚谷氨酸-D-半乳糖酯化衍生物-顺铂复合物是一种有效的具有肝靶向性的抗肿瘤药物,具有潜在的临床应用价值;通过生物发酵获得的γ-聚谷氨酸可用于药物载体,赋予药物新的特点。     

Differential radioprotection of tissues in C3H/J mice by a combination of 5-hydroxy L-tryptophan with 2-aminoethylisothiuronium bromide hydrobromide.

Differential radioprotection between normal tissues and carcinoma was observed in C3H/J mice treated with a combination of 5-hydroxy L-tryptophan (5-HTP, 100 mg/kg) and 2-aminoethylisothiuronium bromide hydrobromide (AET, 20 mg/kg). Protection to normal tissues was judged by LD50(30) and by radiation induced damage to bone marrow(BM) using clonogenic ability of blood forming stem cells (10 day CFUs) as the criteria. Pretreatment with 5-HTP + AET combination 30 min before whole body gamma radiation (WBGR) enhanced the recoveries of the number of blood forming stem cells in BM of irradiated mice after 0, 7th and 10th day of irradiation. LD50(30) for C3H/J mice was 7.3 Gy and the dose modifying factor (DMF) of 5-HTP + AET combination was 1.76. On the contrary, pretreatment with this combination did not protect the mammary carcinoma transplanted in C3H/J mice, when exposed to 80 Gy soft X-rays.     

Naloxone prolongs the survival time of mice treated with neuroblastoma

The effect of naloxone on tumor growth and survival time was studied in mice with neuroblastoma tumors. Daily s.c. injections of either 5, 10, 15, or 20 mg/kg naloxone were initiated either 2 weeks prior to tumor cell inoculations (pre-treated groups) or one week after tumor transplantation (post-treated groups). The S20Y cell line, cloned from A/Jax murine C1300 neuroblastoma, was utilized and each animal was inoculated with 10⁶ cells. All mice in the saline- tumor and naloxone post-treated groups, developed tumors within 3 weeks after tumor cell inoculation. In the naloxone pre-treated groups, 4 of 12 mice exposed to 20 mg/kg, 2 of 12 mice exposed to 15 mg/kg, and 1 of 12 mice exposed to 10 mg/kg, did not develop tumors within the 91-day post-inoculation period. Three animals in the 20 mg/kg naloxone pre-treated group developed tumors between 43 and 63 days after tumor cell inoculation. Tumor dimensions were often reduced in naloxone-treated animals but a dose-response relationship was not found in regard to the magnitude of alterations in tumor size. At the time of death, tumor sizes of control and naloxone-exposed mice were similar. In general, tumor-bearing mice receiving naloxone lived longer than saline-tumor controls, with animals receiving higher drug dosages surviving for the longest time. In contrast to a mean survival time of 27 days for controls, naloxone pre-treated groups had increases in survival times of 25–61%, whereas naloxone post- treated groups exhibited increases of 20–40%. The median day of death for all mice exposed to naloxone was prolonged by 21–75%, occuring 6–21 days after the 28-day median for saline-tumor controls. These results suggest that naloxone, a non-addictive compound, is an effective agent in modulating neoplasia.     

The T-cell receptor: just another hypothesis.

Functional activities of T and B lymphocytes and the kinetics of hematopoietic stem cells were studied in mice with inoculated or spontaneous tumors. The development and growth of the tumor inhibited B cells and helper T cells, while the activity of killer T cells and spleen suppressor cells was markedly enhanced. The processes of stem cell migration from the bone marrow were considerably intensified and altered in tumor-bearing mice. Data were obtained suggesting that helper T cells and killer T cells represent nonidentical compartments within the population of thymus-dependent lymphocytes. Immunosuppression during tumor bearing is probably due to an impairment of T lymphocytes cooperating in immune responses, B-lymphocytes and their precursors.     

T and B lymphocyte migration into syngeneic tumors.

The migration of splenic T and B lymphocytes into syngeneic tumors undergoing immunologic rejection was investigates. Spleen cells were obtained from normal BALC/c mice or BALB/c mice bearing tumors induced by murine sarcoma virus (MSV). Either whole spleen cells or immunoabsorbent purified T and B cells were radiolabeled with sodium chromate-51 and injected i.v. into normal or MSV inducted-tumor bearing syngeneic recipients. Twenty-four hours later the recipient mice were sacrificed and radioactivity was assessed for tumor, contralateral normal muscle, the lymph nodes draining the tumor and contralateral draining lymph nodes, peripheral lymph nodes, spleen, and liver. Both T and B lymphocytes from either normal or MSV tumor-bearing animals show greatly increased migration into the tumor when compared with normal muscle. Migration of T cells from both normal and MSV tumor bearers was 30 times that of migration to normal muscle. B cells from tumor-bearing mice, on the other hand, localized in the tumor itself only 50% as frequently as did B cells from normal animals. In addition, T cells from MSV tumor bearers were found in the highest proportion in the lymph node draining the tumor site. We conclude that T and B lymphocytes from either normal or tumor-bearing mice migrate to a syngeneic tumor undergoing immunologic rejection. In contrast, the migration of both T and B cells from tumor-bearing animals was decreased to the peripheral lymph nodes at the time of maximum tumor growth.     

Normal tissue alloantigens and genetic control of susceptibility to tumors: microcytotoxicity studies on resistant C3Hf and susceptible (A X C3Hf) F1 mice inoculated with transplacentally induced C3Hf lung tumor.

The immune response of mice to a transplacentally induced lung tumor was investigated with the microcytotoxicity (MC) assay. The tumor, originally induced in C3Hf mice, does not grow readily when transplanted to normal syngeneic C3Hf recipients. It grows readily, however, in (A C3Hf)F1 hybrids and in strain C3H mice, which express in their normal lung tissue a component which constitutes a strong lung tumor-associated transplantation antigen (TATA) in C3Hf mice. Both lung tumor-immunized C3Hf and tumor-bearing (A X C3Hf)F1 and C3H mice possessed lymphoid cells reactive against cultured lung tumor cells in the MC assay. Reactivity was also observed against cells cultured from normal lungs of (A X C3Hf)F1 and C3H mice, but not against cells similarly cultured from C3Hf of C57BL/6 mice. Anti-tumor MC was inhibited by serum-blocking factors present in some but not all tumor-bearing and tumor-immunized mice. The MC assay and detection by it of serum-blocking factors does not distinguish the effective anti-C3Hf lung tumor immune response of immunized C3Hf mice from the ineffective immune response of tumor-bearing (A X C3Hf)F1 and C3H mice. Furthermore, in lung tumor-bearing mice cells reactive in the MC assay may be directed against a normal tissue antigen rather than a tumor-associated antigen.     

A majority of mice show long-term expression of a human beta-globin gene after retrovirus transfer into hematopoietic stem cells.

Murine bone marrow was infected with a high-titer retrovirus vector containing the human beta-globin and neomycin phosphotransferase genes. Anemic W/Wv mice were transplanted with infected marrow which in some cases had been exposed to the selective agent G418. Human beta-globin expression was monitored in transplanted animals by using a monoclonal antibody specific for human beta-globin polypeptide, and hematopoietic reconstitution was monitored by using donor and recipient mice which differed in hemoglobin type. In some experiments all transplanted mice expressed the human beta-globin polypeptide for over 4 months, and up to 50% of peripheral erythrocytes contained detectable levels of polypeptide. DNA analysis of transplanted animals revealed that virtually every myeloid cell contained a provirus. Integration site analysis and reconstitution of secondary marrow recipients suggested that every mouse was reconstituted with at least one infected stem cell which had extensive repopulation capability. The ability to consistently transfer an active beta-globin gene into mouse hematopoietic cells improves the feasibility of using these techniques for somatic cell gene therapy in humans.     

骨髓移植小鼠二甲基亚硝胺肝纤维化模型的建立

目的构建绿色荧光蛋白(GFP)标记骨髓细胞的小鼠,并复制其二甲基亚硝胺(DMN)肝纤维化模型。方法 ICR雄性小鼠32只,随机分为正常组6只和移植组26只。移植组接受致死量γ射线照射后,经尾静脉输入GFP转基因小鼠的骨髓细胞;正常组不进行照射和移植,仅尾静脉注射等量生理盐水。两个月后制备血涂片,观察移植组造血重建情况,造血重建动物再分为对照组和造模组,造模组用DMN按每次10mg/kg体重腹腔注射制备肝纤维化模型,隔日一次,正常组和对照组给予等量生理盐水。设染毒后3周和4周两个时间观察点,生化法测定肝功能;Jamall法检测肝组织羟脯氨酸含量;HE染色及天狼猩红染色观察肝组织炎症、坏死及纤维组织增生情况;GFP免疫荧光组织化学观察骨髓源性细胞在肝脏的归巢特点。结果骨髓移植两个月后,移植组外周血中出现满视野GFP+细胞。与正常组比较,两个时间观察点造模组肝功能(ALT、AST、Alb及T.Bil)均明显异常(P<0.05),肝组织羟脯氨酸含量显著增高(P<0.05),造模3周末肝组织出血性坏死,有炎性细胞浸润,但尚未形成完全的纤维间隔;造模4周末肝组织炎症、坏死程度加重,可见完全的纤维间隔,在DMN造模动物肝组...     

Parathyroid hormone (PTH) and hematopoiesis: new support for some old observations

Forty-seven years ago, the parathyroid hormone (PTH) in one injection of Lilly's old bovine parathyroid extract, PTE, was found to greatly increase the 30-day survival of heavily X-irradiated rats when given from 18 h before to as long as 3 h after irradiation but no later. This was the first indication that PTH might stimulate hematopoiesis. Recent studies have confirmed the relation between PTH and hematopoiesis by showing that hPTH-(1-34)OH increases the size of the hematopoietic stem cell pool in mice. The peptide operates through a cyclic AMP-mediated burst of Jagged 1 production in osteoblastic cells lining the stem cells' niches on trabecular bone surfaces. The osteoblastic cells' Jagged 1 increases the hematopoietic stem cell pool by activating Notch receptors on attached stem cells. PTH-triggered cyclic AMP signals also directly stimulate the proliferation of the hematopoietic stem cells. However, the single PTH injection in the early experiments using PTE probably increased the survival of irradiated rats mainly by preventing the damaged hematopoietic progenitors from irreversibly initiating self-destructive apoptogenesis during the first 5 h after irradiation. It has also been shown that several daily injections of hPTH-(1-34)OH enable lethally irradiated mice to survive by stimulating the growth of transplanted normal bone marrow cells. If the osteogenic PTHs currently entering or on the verge of entering the market for treating osteoporosis can also drive hematopoiesis in humans as well as rodents, they could be potent tools for reducing the damage inflicted on bone marrow by cytotoxic cancer chemotherapeutic drugs and ionizing radiation.     

Generation of murine stromal cell lines supporting hematopoietic stem cell proliferation by use of recombinant retrovirus vectors encoding simian virus 40 large T antigen.

The bone marrow is a complex microenvironment made up of multiple cell types which appears to play an important role in the maintenance of hematopoietic stem cell self-renewal and proliferation. We used murine long-term marrow cultures and a defective recombinant retrovirus vector containing the simian virus 40 large T antigen to immortalize marrow stromal cells which can support hematopoiesis in vitro for up to 5 weeks. Such cloned cell lines differentially supported stem cells which, when transplanted, allowed survival of lethally irradiated mice, formed hematopoietic spleen colonies in vivo, and stimulated lymphocyte proliferation in vitro. Molecular and functional analyses of these cell lines did not demonstrate the production of any growth factors known to support the proliferation of primitive hematopoietic stem cells. All cell lines examined produced macrophage colony-stimulating factor. The use of immortalizing retrovirus vectors may allow determination of unique cellular proteins important in hematopoietic stem cell proliferation by the systematic comparison of stromal cells derived from a variety of murine tissues.     

Chemoprotective effects of recombinant human IL-1 alpha in cyclophosphamide-treated normal and tumor-bearing mice. Protection from acute toxicity, hematologic effects, development of late mortality, and enhanced therapeutic efficacy

In this study, recombinant human IL-1 alpha (rhIL-1 alpha) was used to protect normal and tumor-bearing BALB/c mice from the acute toxicity caused by lethal doses of cyclophosphamide (Cy) and 5-fluorouracil. Pretreatment of mice for 7 days with 10,000 U/day of rhIL-1 alpha protected 70 to 100% of mice from the acute death induced by lethal doses of both Cy (380 mg/kg) and 5-fluorouracil (250 mg/kg). In contrast, post-treatment of mice with single or multiple doses of rhIL-1 alpha was not chemoprotective. Pretreatment of mice with rhIL-1 alpha increased the acute LD90 of Cy from 380 mg/kg to greater than 500 mg/kg in normal mice, an LD90 dose-modifying effect of at least 1.25, was accompanied by a more rapid recovery from neutropenia and a less severe reduction in the number of bone marrow single lineage monocyte, myeloid, or myelomonocytic colonies. Some of the mice (10 to 50%) that were successfully protected by pretreatment with rhIL-1 alpha died after day 50. These mice consistently presented with extensive pulmonary inflammation and fibrosis at death. Mice bearing murine renal cancer (Renca) were also protected from the acute toxic effects of Cy (450 mg/kg) by pretreatment with rhIL-1 alpha. Renca-bearing mice pretreated with rhIL-1 alpha and either sublethal (300 mg/kg) or lethal (450 mg/kg) doses of Cy exhibited enhanced survival times over those of untreated Renca-bearing mice. Interestingly, the cause of death in Renca-bearing mice that ultimately failed treatment with rhIL-1 alpha plus 300 mg/kg Cy was recurrent tumor, whereas most mice treated with rhIL-1 alpha plus 450 mg/kg Cy had no detectable tumor, although several died from late pulmonary inflammation and fibrosis. Thus, the dose escalation of Cy in rhIL-1 alpha-pretreated mice results in greater antitumor effects of Cy. However, the dose escalation of some cytotoxic agents allowed by the use of myelostimulatory agents can result in late fatal complications not detected in acute toxicity testing.     

Significance of host heparanase in promoting tumor growth and metastasis

Heparanase, the sole heparan sulfate degrading endoglycosidase, regulates multiple biological activities that enhance tumor growth, angiogenesis and metastasis. Much of the impact of heparanase on tumor progression is related to its function in mediating tumor-host crosstalk, priming the tumor microenvironment to better support tumor growth and metastasis. We have utilized mice over-expressing (Hpa-tg) heparanase to reveal the role of host heparanase in tumor initiation, growth and metastasis. While in wild type mice tumor development in response to DMBA carcinogenesis was restricted to the mammary gland, Hpa-tg mice developed tumors also in their lungs and liver, associating with reduced survival of the tumor-bearing mice. Consistently, xenograft tumors (lymphoma, melanoma, lung carcinoma, pancreatic carcinoma) transplanted in Hpa-tg mice exhibited accelerated tumor growth and shorter survival of the tumor-bearing mice compared with wild type mice. Hpa-tg mice were also more prone to the development of metastases following intravenous or subcutaneous injection of tumor cells. In some models, the growth advantage was associated with infiltration of heparanase-high host cells into the tumors. However, in other models, heparanase-high host cells were not detected in the primary tumor, implying that the growth advantage in Hpa-tg mice is due to systemic factors. Indeed, we found that plasma from Hpa-tg mice enhanced tumor cell migration and invasion attributed to increased levels of pro-tumorigenic factors (i.e., RANKL, SPARC, MIP-2) in the plasma of Hpa-Tg vs. wild type mice. Furthermore, tumor aggressiveness and short survival time were demonstrated in wild type mice transplanted with bone marrow derived from Hpa-tg but not wild type mice. These results were attributed, among other factors, to upregulation of pro-tumorigenic (i.e., IL35⁺) and downregulation of anti-tumorigenic (i.e., IFN-γ⁺) T-cell subpopulations in the spleen, lymph nodes and blood of Hpa-tg vs. wild type mice and their increased infiltration into the primary tumor. Collectively, our results emphasize the significance of host heparanase in mediating the pro-tumorigenic and pro-metastatic interactions between the tumor cells and the host tumor microenvironment, immune cells and systemic factors.     

Evaluation of Tempol Radioprotection in a Murine Tumor Model

Tempol, a stable nitroxide free radical compound, is an in vitro and in vivo radioprotector. Previous studies have shown that Tempol protects C3H mice against whole-body radiation-induced bone marrow failure. In this study, the radioprotection of tumor tissue was evaluated. RIF-1 tumor cells were implanted in female C3H mice 10 d prior to radiation. Groups of mice were injected intraperitoneally with Tempol (275 mg/kg) or PBS followed 10 min later by a single dose of radiation to the tumor bed. Tumor growth curves generated after 10 and 33.3 Gy doses of radiation showed no difference in growth between the Tempol- and PBS-treated animals. A full radiation dose-response experiment revealed a tumor control dose in 50% of the animals in 30 d (TCD_50/30) value of 36.7 Gy for Tempol-treated mice and 41.8 Gy for saline-treated mice suggesting no protection of the RIF-1 tumor by Tempol. Tumor pharmacokinetics were done to determine why Tempol differentially protected bone marrow and not tumor cells. Differential reduction of Tempol in the RIF-1 tumor and bone marrow was evaluated with EPR spectroscopy 10, 20, and 30 min after injection. Bioreduction of Tempol to its corresponding hydroxylamine (which is not a radioprotector) occurred to a greater extent in RIF-1 tumor cells compared to bone marrow. We conclude that the differences in radioprotection may result from enhanced intratumor bioreduction of Tempol to its nonradioprotective hydroxylamine analogue. The nitroxides as a class of compounds may provide a means to exploit the redox differences between normal tissues and tumors. © 1997 Elsevier Science Inc.     

Modulation of the immune response to tumors by a novel synthetic compound, (4R)-3-benzoyl-N-[(1R)-1-phenylethyl]-4-thiazolidinecarboxamide (RS-0481)

Summary RS-0481, (4R)-3-benzoyl-N-[(1R)-phenylethyl]-4-thiazolidinecarboxamide, is a compound that can re-establish the function of certain lymphoid cell populations impaired by the presence of a growing tumor in an animal. The compound markedly augmented the tumorspecific cytotoxic T lymphocytes,Tdth (delayed-type hypersensitivity T cells), and the nonspecific lymphokine-activated-killer-cell-like cell responses. It also enhanced the tumor-inhibitory effect of macrophages in tumor-bearing mice, but not in normal mice, indicating that it enhances the antitumor immune responses. Lymphocytes from RS-0481-treated tumor-bearing mice released significantly higher amounts of macrophage-activating factor(s) (MAF) and interleukin-2(IL-2)-like factors in culture compared with lymphocytes from untreated animals. Also, sera from treated tumor bearers showed elevated colony-stimulating factor (CSF) activity. Although the compound did not influence the factor-producing activity in mice without tumor, it enhanced the responsiveness of their bone marrow cells, T cells, and macrophages to CSF, IL-2, and MAF. It seems therefore possible that the compound enhances the responsiveness of immunocompetent cells to cytokines, resulting in a marked augmentation of antitumor T cell responses in tumor-bearing mice. Consistently it inhibited the development of lymph node metastasis of transplanted X5563 plasmacytoma, and we showed that T cells play a decisive role in this inhibition. The compound also counteracted the development of suppressor T cell activity in the spleen of tumor-bearing mice.