Saikosaponin-d regulates angiogenesis in idiopathic pulmonary fibrosis through angiopoietin/Tie-2 pathway.

OBJECTIVE: Idiopathic pulmonary fibrosis (IPF) is considered as a chronic interstitial lung disease with underlying mechanism of IPF remaining unclear, while there are no definitive treatment options. In recent years, scientists have gradually paid attention to the influence of angiogenesis on IPF. Because IPF is a progressive with microvascular remodeling disorder, scientists have postulated that angiogenesis may also be one of the initiating and contributing factors of the disease. Bupleurum is a common natural Chinese herbal medicine with antibacterial, anti-inflammatory, anti-tumor and other pharmacological effects. As the most important active monomer of Bupleurum, Saikosaponin-d (SSd) is a new discovery with anti-pulmonary fibrosis (PF) activity. This study attempts to investigate the role of SSd in the interference of PF through regulation of angiogenesis in IPF through Angiopoietin (Angpt) /Tie receptor 2 (Tie2) pathway. METHODS: Randomly, we allocated C57BL/6 mice into four groups (n = 20 in each group). Afterwards, establishment of IPF model was accomplished via intratracheal administration of bleomycin (BLM, 5 mg/kg), while corresponding drug intervention was given accordingly. On 3rd, 7th, 14th and 28th days after modeling, we performed histopathological examination through staining. Meanwhile, immunohistochemistry (IHC) of PF and the expression of related factors were observed, while Ang/Tie2 pathway was assessed by ELISA with the effect of SSd on angiogenesis related proteins in IPF being explored with IHC and Western Blot technique. RESULTS: Our results showed that SSd could reduce inflammation and PF levels in lung tissue of experimental mice, while levels of angiogenesis-related factors, namely Tie-2, Ang-1 and ANGPT2 (Ang-2), fibrosis- associated factors like Alpha-smooth muscle actin (α-SMA), collagen-I and hydroxyproline in SSd and dexamethasone (DXM) mice were significantly reduced at each time point compared to BLM (p < 0.01). Additionally, we discovered substantial decreased expressions of Ang-1, Ang-2, Tie-2, α-SMA and collagen-I at protein level in SSd and DXM mice at each time point compared to BLM (p < 0.05). Besides, insignificant differences were observed between SSd and DXM groups (p > 0.05). CONCLUSION: This study has demonstrated that SSd could down-regulate the expression of ANG-1, Ang-2 and Tie2 in the Ang/Tie2 pathway, and may reduce lung inflammation and PF in BLM-induced mice via inhibition of angiogenesis.

Total sesquiterpenoids from Eupatorium lindleyanum DC. attenuate bleomycin-induced lung fibrosis by suppressing myofibroblast transition.

Eupatorium lindleyanum DC. has been used as a functional food in China for a long time. However, the antifibrotic activity of total sesquiterpenoids from Eupatorium lindleyanum DC. (TS-EL) is still unknown. In this study, we discovered that TS-EL reduced the increase in α-smooth muscle actin (α-SMA), type I collagen and fibronectin content, the formation of cell filaments and collagen gel contraction in transforming growth factor-ß1-stimulated human lung fibroblasts. Intriguingly, TS-EL did not change the phosphorylation of Smad2/3 and Erk1/2. TS-EL decreased the levels of serum response factor (SRF), a critical transcription factor of α-SMA, and SRF knockdown alleviated the transition of lung myofibroblasts. Furthermore, TS-EL significantly attenuated bleomycin (BLM)-induced lung pathology and collagen deposition and reduced the levels of two profibrotic markers, total lung hydroxyproline and α-SMA. TS-EL also decreased the levels of SRF protein expression in BLM-induced mice. These results suggested that TS-EL attenuates pulmonary fibrosis by inhibiting myofibroblast transition via the downregulation of SRF.

Efficacy of Qingfei oral liquid for idiopathic pulmonary fibrosis in rats and related network pharmacology study.

To investigate the therapeutic effect and mechanism of Qingfei oral liquid in idiopathic pulmonary fibrosis. Seventy-two male SD rats were divided into control group, model group, pirofenidone group and Qingfei group with 18 animals in each group. The idiopathic pulmonary fibrosis was induced in last three groups by intratracheal injection of bleomycin; pirofenidone group was given oral administration of pirofenidone b.i.d for 21 d, and Qingfei group was given Qingfei oral liquid 3.6 mL/kg q.d for Lung tissues were obtained for HE staining, Masson staining and transforming growth factor (TGF)-ß immunohistochemical staining. Superoxide dismutase (SOD), malondialdehyde (MDA) and glutathione (GSH) were detected in tissue homogenates. The BATMAN-TCM database was used to retrieve the chemical components and their corresponding targets of Qingfei oral solution by network pharmacology method, and then the component-target-disease network diagram was constructed. Finally, the pathway enrichment analysis was carried out to explore the molecular mechanism of Qingfei oral liquid against idiopathic fibrosis. Histopathology results showed that Qingfei oral liquid had a similar relieving effect on pulmonary fibrosis as the positive drug pirfenidone; TGF-ß secretion had a significant reduction in lung tissues of Qingfei group; and Qingfei oral liquid had better regulatory effect on SOD, MDA and GSH than pirfenidone. The results of component-target-disease network and pathway enrichment analysis showed that the related molecular pathways were concentrated in inflammation, extracellular matrix and cytokines. Qingfei oral liquid has a good therapeutic effect on idiopathic pulmonary fibrosis in rats via regulation of inflammation, extracellular matrix and cytokines.

Evaluating the protective role of Deglycyrrhizinated licorice root supplement on bleomycin induced pulmonary oxidative damage.

The goal of this study was to investigate the protective effect of licorice supplements in a rat model of Bleomycin-induced lung oxidative damage over a duration of one month. The rats were randomly divided into six groups (n = 10 per group). Control group; Bleomycin group (B): rats were IP injected with bleomycin 5 mg/kg twice weekly. Licorice group (L): rats received orally 300 mg/kg licorice extract. Bleomycin and a low dose of Licorice group (BLLG): rats received orally 75 mg/kg licorice daily and injected as the B group. Bleomycin and a middle dose of Licorice group (BMLG): rats received orally 150 mg/kg licorice daily and injected as the Bleomycin group. Bleomycin and a high dose of Licorice group (BHLG): rats received orally 300 mg/kg licorice daily and injected as the Bleomycin group. Treatment with Bleomycin induced inflammation and oxidative damage to the lungs expressed in the disturbance of the measured parameters in the blood serum, the lung tissue, and the broncholavage fluid. In addition to the decreased expression of superoxide dismutase (SOD), glutathione peroxidase (GSH-PX), and catalase (CAT) in the lung tissues. Bleomycin caused deformative changes in the histopathological and cellular examination of the lungs especially in the alveolar cells and the interstitial space. On the other hand, treated the bleomycin group with different doses of licorice supplement activates the antioxidant defense mechanism and attenuates the oxidative damage and damage induced to the lung. In conclusion, Deglycyrrhizinated licorice root supplement provided strong antioxidant and protective effects on Bleomycin-induced lung damage.

Gigantic cavernous hemangioma of the liver treated by intra-arterial embolization with pingyangmycin-lipiodol emulsion: a multi-center study.

PURPOSE: To evaluate the therapeutic effect and safety of pingyangmycin-lipiodol emulsion (PLE) intra-arterial embolization for treating gigantic cavernous hemangioma of the liver (CHL). METHODS: Three hospitals (Nanfang Hospital, Inner Mongolia Autonomous Region's Hospital and Huai He Hospital) participated in the study during 1997-2001. A total of 98 patients with CHL were embolized with PLE via the hepatic artery. The therapeutic effects including changes in tumor diameter, symptomatic improvement and occurrence of complications were evaluated for a period of 12 months after the procedure. RESULTS: The tumor diameters decreased significantly from 9.7 +/- 2.3 cm to 5.6 +/- 1.6 cm 6 months after the treatment ( P < 0.01), and then to 3.0 +/- 1.2 cm at 12 months ( P < 0.01). Transient impairment of liver function was found in 77 cases after embolization, 69 cases of which returned to normal in 2 weeks, and the other eight cases of which recovered 1 month later. The clinical symptoms were significantly relieved in all 53 symptomatic patients. Persistent pain in the hepatic region was found in two cases, and these two patients resorted to surgery eventually. CONCLUSION: Intra-arterial PLE embolization proves to be effective and safe in treating patients with CHL.

Sequence-specific changes in the metal site of ferric bleomycin induced by the binding of DNA.

The binding of the iron complex of the antineoplastic glycopeptide bleomycin A2 (Fe-BLM) to calf thymus DNA and the self-complementary oligonucleotides d(CGCGCG) and d(ATATAT) has been studied using optical, EPR, and resonance Raman spectroscopies. An increase in the intensity of the bands at 365 and 384 nm is observed in the optical spectrum of Fe(III)-BLM when the drug binds to either oligonucleotide. However, in the presence of phosphate, this increase is observed only with d(CGCGCG) and not with d(ATATAT). In addition, the gmax feature in the EPR spectrum of low spin Fe(III)-BLM is narrowed in a way suggesting a reduction of possible conformers that the drug can achieve when it is bound to d(CGCGCG) or to calf thymus DNA but not when bound to d(ATATAT). When Fe(III)-BLM is bound to d(CGCGCG), changes in the resonance Raman spectrum of the metal drug complex suggest conformational changes in three of the ligands to iron: the beta-hydroxyhistidyl amide, the pyrimidine, and the axial hydroxide. In addition, the Fe-OH band undergoes narrowing, again consistent, with the reduction of conformers of the drug. No such resonance Raman changes are observed upon binding to d(ATATAT). The changes in the pyrimidine modes upon binding d(CGCGCG) to the drug are consistent with a recently proposed model (Wu, W., Vanderwall, D. E., Turner, C. J., Kozarich, J. W., and Stubbe, J. (1996) J. Am. Chem. Soc. 118, 1281-1294) of DNA recognition by activated bleomycin, HOO-Fe(III)-BLM, in which the pyrimidine moiety of the drug is important for the preferential cleavage of 5'-GpPy-3' sequences.

Ascorbate both activates and inactivates bleomycin by free radical generation.

The chemotherapeutic agent bleomycin (BLM) is activated by reducing agents to break isolated DNA. Paradoxically, these same reducing agents protect cellular DNA from BLM damage. To resolve this paradox, we have examined the reaction of FeIIIBLM with DNA in the presence of ascorbate. As expected, ascorbate augments FeIIIBLM-induced DNA damage. However, when ascorbate is added to FeIIIBLM prior to exposure to DNA, a redox-inactive BLM is produced in a reaction that generates the ascorbyl radical. This reaction occurs in both ascorbate-supplemented buffer and unsupplemented plasma. In buffered solution, this reaction was found to be stoichiometric; for each mole of BLM present, 6.9 mol of ascorbate was oxidized and 4.7 mol of oxygen was consumed. Iron was found to serve only as a catalyst for the reaction. These data suggest that both activation of BLM and the generation of redox-inactive BLM occur via the same reaction and that BLM-induced DNA damage depends upon BLM reaching DNA prior to its interaction with reducing agents.

Plasma ascorbate levels and inhibition of the antioxidant activity of caeruloplasmin.

1. The copper-containing protein caeruloplasmin has several oxidase activities. 2. Its ability to catalyse the oxidation of ferrous ions to the ferric state (ferroxidase activity) makes it an important antioxidant in vivo. 3. Recent reports have suggested that oral supplementation with vitamin C can inhibit the oxidase activities of caeruloplasmin. 4. As expected, damage to DNA and membrane lipids was stimulated by mixtures of iron salt and ascorbate, and this damage could be inhibited by caeruloplasmin provided the molar ratio of ascorbate to caeruloplasmin was kept sufficiently low. 5. When the molar ratio of ascorbate to caeruloplasmin was greater than 200 substantial loss of ferroxidase antioxidant activity occurred. 6. It is unlikely, however, that oral supplementation with vitamin C can raise plasma levels sufficiently to inhibit caeruloplasmin activity in vivo.

[Experimental research following intratumor bleomycin use in the nude mouse model of oral mucosa cancer and the clinical pilot study]. / Experimentelle Untersuchungen nach intratumoraler Bleomycin-Applikation am nude-Maus-Modell des Mundschleimhautkarzinoms und klinische Pilotstudie.

Experiments concerning the kinetics and bio-distribution of 57Co-Bleomycin in three different lines of the squamous cell carcinoma of the oral cavity in a nude-mouse model yielded a more than tenfold higher concentration after a single intratumoral (i.t.) application of an aqueous solution of 57Co-labelled Bleomycin in spite of a remarkable radioactivity-wash in the tumor up to 48 hours post applicationem, compared to the intravenous (i.v.) injection. For the investigation of the radiopharmacon within the tumor we used the macro-autoradiographic method. The xenografts have been removed and cut (5 micrograms) 1;5; 24, and 48 hours after the Bleomycin administration, respectively. These preparations have been covered with an autoradiographic film and exposed for about three weeks. After this an inhomogenous distribution of the radioactive nuclide was produced within the tumor, and a particularly high activity-concentration could be demonstrated in the necrotic tumor areas. The results obtained from the animal experiments have been corroborated by a pilot-study consisting of i.t. Bleomycin application in nine patients with a carcinoma of the oral mucosa.

Effect of adjuvants on the rectal absorption and lymphatic uptake of pepleomycin in rats.

The rectal absorption of pepleomycin sulfate (PEPS) in rats was increased significantly by the coadministration with each of diclofenac (DC), sodium 5-methoxysalicylate (5-MSA) and phenylalanine enamine of ethylacetoacetate (Enamine). 5-MSA increased the lymphatic uptake of PEPS after rectal administration while DC and Enamine did not. The mechanism behind the enhancing action of 5-MSA on the lymphatic uptake of PEPS may be due to the suppressing action of 5-MSA on the vascular permeability to PEPS. DC increased the vascular permeability to PEPS but Enamine did not affect it. Findings obtained in this study may indicate that adjuvant used acts independently at the rectal mucosal membrane and at the vascular membrane for membrane for membrane permeability to PEPS.

Clinical pharmacokinetics of commonly used anticancer drugs.

The quantitative aspects of drug disposition in man of the commonly used antineoplastic agents, including cyclophosphamide, the nitrosoureas, cisplatin, methotrexate, cytarabine, 5-fluorouracil, doxorubicin, daunorubicin, bleomycin, vincristine, vinblastine, and vindesine are reviewed. Although the pharmacokinetic behaviour of these drugs has been adequately described in man, the chemical reactivity, the complexity of metabolism and disposition, the lack of simple, rapid and sensitive assays to measure plasma concentration, and the lack of defined therapeutic and toxic plasma concentrations have limited the application of routine drug monitoring in clinical oncology. With the exception of high dose methotrexate, drug doses and administration schedules remain empirical with a standard starting dose and subsequent dosage modifications determined by ensuing drug toxicities. However, many of the pharmacological characteristics of the drugs, such as their low therapeutic index, potentially life-threatening toxicities and wide individual variability in drug disposition, necessitate pharmacological monitoring. Comprehensive pharmacokinetic analysis of new and established antineoplastic agents does play a role in defining dosage, administration schedule, route of administration, and dosage modification in the presence of organ dysfunction. Consideration of the kinetics of these drugs in planning treatment regimens could lead to more rational, safer and possibly more efficacious use.

[Studies on organ distribution, absorption and excretion of pepleomycin sulfate (NK631) (author's transl)].

1. Organ distribution of pepleomycin (NK631) in mice and rats was studied. NK631 was found at higher levels than bleomycin (BLM) in skin, lung, stomach, solid tumor, etc. in mice and rats. Furthermore NK631 was detected in the mesenteric and lumbar lymph node, esophagus and prostate in rats and also distributed at about twice as high levels as BLM in the AH109A hepatoma cell-metastasized lymph nodes. 2. For the elucidation of reason on low pulmonary toxicity of NK631 which is in spite of 1.5 times highly distribution in lung compared with BLM, inactivation of various BLMs by high molecular fraction of lung of mice and rats was determined. The order of inactivation rate of various BLMs in lung was as follows: BLM-M5196 greater than NK631 greater than BLM greater than BLM-HPE. There is an encouraging coincidence between index of pulmonary fibrosis in mice and inactivating rate in lung. 3. A comparative study on the serum level and urinary excretion of NK631 and BLM was performed in dogs. The blood level and urinary excretion rate of both drugs were almost similar. 4. The blood levels of NK631 were comparable to those of BLM in cancer patients.

Enhancement of the cancer chemotherapeutic effect by anticancer agents in the form of fat emulsion.

Utilizing the lipid-adsorbing ability of lymphatic capillaries, anticancer agents were given in the form of fat emulsion in order to deliver them to regional lymph nodes. The emulsion, in which the drug solution is contained as the innermost phase, yielded high drug concentration in the lymphatic system. Intratumoral injection of emulsified anticancer agent resulted in significantly prolonged retension of the drug within the tumor tissue. Therapeutic experiments of the emulsion also disclosed remarkable tumor reduction and cure rate as compared with aqueous solution of drugs. Oral administration of emulsified 5-Fluorouracil (5-FU) was also attempted for stomach cancer. With 5-FU, the maximum concentration of drug in thoracic lymph and stomach was greater when administered as an emulsion than as an aqueous solution, and a high concentration persisted longer. As a clinical trial of the emulsion method, eight patients with inoperable malignant growth were injected locally with emulsified anticancer agents and 121 patients were given 5-FU emulsion orally. From the clinical and histological findings, it was thought that the emulsion enhanced the chemotherapeutic effect of the anticancer agent on lymph node metastasis.