This study investigated the effect of TNP-470 in combination with carmustine (BCNU) on the growth of subcutaneously implanted human glioblastoma xenografts in nude mice. Human glioblastoma U-251 cells (1×10(7)) were injected into 24 nude mice subcutaneously. The tumor-bearing mice were randomly divided into 4 groups on the seventh day following tumor implantation: TNP-470 group, in which TNP-470 was given 30 mg/kg subcutaneously every other day 7 times; BCNU group, in which 20 mg/kg BCNU were injected into peritoneal cavity per 4 days 3 times; TNP-470 plus BCNU group, in which TNP-470 and BCNU were coadministered in the same manner as in the TNP-470 group and the BCNU group; control group, in which the mice were given 0.2 mL of the mixture including 3% ethanol, 5% acacia and 0.9% saline subcutaneously every other day 7 times. The tumor size and weights were measured. The tumor microvessel density (MVD) was determined by immunostaining by using goat-anti-mouse polyclonal antibody CD105. The results showed that on the 21th day following treatment, the volume of xenografts in the TNP-470 plus BCNU group was (108.93±17.63)mm(3), markedly lower than that in the TNP-470 group [(576.10±114.29)mm(3)] and the BCNU group [(473.01±48.04)mm(3)] (both P<0.01). And the xenograft volume in these 3 treatment groups was even much lower than that in the control group [(1512.61±470.25) mm(3)] (all P<0.01). There was no significant difference in the volume of xenografts between the TNP-470 group and the BCNU group (P>0.05). The inhibition rate of the tumor growth in the TNP-470 plus BCNU group was (92.80±11.37)%, notably higher than that in the TNP-470 group [(61.91±6.29)%] and the BCNU group [(68.73±9.65)%] (both P<0.01) on the 21th day following treatment. There was no significant difference in the inhibition rate of tumor growth between the TNP-470 group and the BCNU group (P>0.05). The MVD of xenografts in the TNP-470 plus BCNU group was decreased significantly as compared with that in the TNP-470 group or the BCNU group (both P<0.05). The MVD of xenografts in the 3 treatment groups was markedly reduced as compared with that in the control group (all P<0.05). No significant changes in weights were observed before and after the treatment in each group (all P>0.05). It was concluded that the combination of TNP-470 and BCNU can significantly inhibit the growth of human glioblastoma xenografts in nude mice without evident side effects.

This study established superparamagnetic iron oxide (SPIO)-labeled nerve growth factor-beta (NGF-beta) gene-modified spinal cord-derived neural stem cells (NSCs). The E14 rat embryonic spinal cord-derived NSCs were isolated and cultured. The cells of the third passage were transfected with plasmid pcDNA3-hNGFbeta by using FuGENE HD transfection reagent. The expression of NGF-beta was measured by immunocytochemistry and Western blotting. The positive clones were selected, allowed to proliferate and then labeled with SPIO, which was mediated by FuGENE HD transfection reagent. Prussian blue staining and transmission electron microscopy (TEM) were used to identify the SPIO particles in the cells. The distinctive markers for stem cells (nestin), neuron (beta-III-tubulin), oligodendrocyte (CNPase) and astrocyte (GFAP) were employed to evaluate the differentiation ability of the labeled cells. The immunocytochemistry and western blotting showed that NGF-beta was expressed in spinal cord-derived NSCs. Prussian blue staining indicated that numerous blue-stained particles appeared in the cytoplasma of the labeled cells. TEM showed that SPIO particles were found in vacuolar structures of different sizes and the cytoplasma. The immunocytochemistry demonstrated that the labeled cells were nestin-positive. After differentiation, the cells expressed beta-III-tubulin, CNPase and GFAP. It was concluded that the SPIO-labeled NGF-beta gene-modified spinal cord-derived NSC were successfully established, which are multipotent and capable of self-renewal.
Cells, Cultured ; Dextrans/*diagnostic use ; Embryo, Mammalian ; Magnetic Resonance Imaging ; Magnetics ; Magnetite Nanoparticles/*diagnostic use ; Nerve Growth Factor/*genetics ; Nerve Growth Factor/pharmacology ; Neural Stem Cells/*cytology ; Spinal Cord/*cytology ; Transfection

This study established superparamagnetic iron oxide (SPIO)-labeled nerve growth factor-β (NGF-β) gene-modified spinal cord-derived neural stem cells (NSCs).The E14 rat embryonic spinal cord-derived NSCs were isolated and cultured.The cells of the third passage were transfected with plasmid pcDNA3-hNGFβ by using FuGENE HD transfection reagent.The expression of NGF-β was measured by immunocytochemistry and Western blotting.The positive clones were selected,allowed to proliferate and then labeled with SPIO,which was mediated by FuGENE HD transfection reagent.Prussian blue staining and transmission electron microscopy (TEM) were used to identify the SPIO particles in the cells.The distinctive markers for stem cells (nestin),neuron (β-Ⅲ-tubulin),oligodendrocyte (CNPase) and astrocyte (GFAP) were employed to evaluate the differentiation ability of the labeled cells.The immunocytochemistry and western blotting showed that NGF-β was expressed in spinal cord-derived NSCs.Prussian blue staining indicated that numerous blue-stained particles appeared in the cytoplasma of the labeled cells.TEM showed that SPIO particles were found in vacuolar structures of different sizes and the cytoplasma.The immunocytochemistry demonstrated that the labeled cells were nestin-positive.After differentiation,the cells expressed β-Ⅲ-tubulin,CNPase and GFAE It was concluded that the SPIO-labeled NGF-β gene-modified spinal cord-derived NSC were successfully established,which are multipotent and capable of self-renewal.

This study investigated the effect of TNP-470 in combination with carmustine (BCNU) on the growth of subcutaneously implanted human glioblastoma xenografts in nude mice. Human glioblastoma U-251 cells (1×10(7)) were injected into 24 nude mice subcutaneously. The tumor-bearing mice were randomly divided into 4 groups on the seventh day following tumor implantation: TNP-470 group, in which TNP-470 was given 30 mg/kg subcutaneously every other day 7 times; BCNU group, in which 20 mg/kg BCNU were injected into peritoneal cavity per 4 days 3 times; TNP-470 plus BCNU group, in which TNP-470 and BCNU were coadministered in the same manner as in the TNP-470 group and the BCNU group; control group, in which the mice were given 0.2 mL of the mixture including 3% ethanol, 5% acacia and 0.9% saline subcutaneously every other day 7 times. The tumor size and weights were measured. The tumor microvessel density (MVD) was determined by immunostaining by using goat-anti-mouse polyclonal antibody CD105. The results showed that on the 21th day following treatment, the volume of xenografts in the TNP-470 plus BCNU group was (108.93±17.63)mm(3), markedly lower than that in the TNP-470 group [(576.10±114.29)mm(3)] and the BCNU group [(473.01±48.04)mm(3)] (both P<0.01). And the xenograft volume in these 3 treatment groups was even much lower than that in the control group [(1512.61±470.25) mm(3)] (all P<0.01). There was no significant difference in the volume of xenografts between the TNP-470 group and the BCNU group (P>0.05). The inhibition rate of the tumor growth in the TNP-470 plus BCNU group was (92.80±11.37)%, notably higher than that in the TNP-470 group [(61.91±6.29)%] and the BCNU group [(68.73±9.65)%] (both P<0.01) on the 21th day following treatment. There was no significant difference in the inhibition rate of tumor growth between the TNP-470 group and the BCNU group (P>0.05). The MVD of xenografts in the TNP-470 plus BCNU group was decreased significantly as compared with that in the TNP-470 group or the BCNU group (both P<0.05). The MVD of xenografts in the 3 treatment groups was markedly reduced as compared with that in the control group (all P<0.05). No significant changes in weights were observed before and after the treatment in each group (all P>0.05). It was concluded that the combination of TNP-470 and BCNU can significantly inhibit the growth of human glioblastoma xenografts in nude mice without evident side effects.
Angiogenesis Inhibitors ; administration & dosage ; Animals ; Antibiotics, Antineoplastic ; administration & dosage ; Antineoplastic Agents, Alkylating ; administration & dosage ; Antineoplastic Combined Chemotherapy Protocols ; therapeutic use ; Brain Neoplasms ; drug therapy ; Carmustine ; administration & dosage ; Cell Line, Tumor ; Cyclohexanes ; administration & dosage ; Female ; Glioblastoma ; drug therapy ; Humans ; Mice ; Mice, Inbred BALB C ; Mice, Nude ; Sesquiterpenes ; administration & dosage ; Xenograft Model Antitumor Assays

ObjectiveBy comparing the difference of volatile components of the decoction pieces before and after being processed by braising method of Jianchangbang and steaming method included in the 2020 edition of Chinese Pharmacopoeia， the influence of processing methods on the flavor formation of Polygoni Multiflori Radix （PMR） was compared. MethodHeadspace-gas chromatography-mass spectrometry （HS-GC-MS） was used to detect the volatile components of 30 batches of PMR samples from 3 origins with 3 processing methods. The GC was performed under programmed temperature （starting temperature of 40 ℃， rising to 150 ℃ at 5 ℃·min^-1， and then rising to 195 ℃ at 10 ℃·min^-1） with high purity helium as carrier gas and the split ratio of 10∶1. Mass spectrometry conditions were electron impact ion source （EI） and the detection range of m/z 50-650， the peak area normalization method was used to calculate the relative mass fraction of each component. The chromaticity values of different processed products were measured by a precision colorimeter， the relationship between chromaticity values and relative contents of volatile components was investigated by OriginPro 2021， principal component analysis （PCA） and orthogonal partial least squares-discriminant analysis （OPLS-DA） were performed on the sample data by SIMCA14.1. The differential components of different processed products of PMR were screened according to the principle of variable importance in the projection （VIP） value>1.5， and the material basis of different odor formation of PMR and its processed products was explored. ResultA total of 59 volatile components were identified， among which 34 were raw products， 33 were braised products， and 27 were steamed products. PCA and OPLS-DA results showed that there were significant differences between the three， but there was no significant difference between samples from different origins of the same processing method. Color parameters of a^*， b^*， E^*ab had no significant correlation with contents of volatile components， while L^* was negatively correlated with contents of 2-methyl-2-butenal， 2-methyltetrahydrofuran-3-one and 2，3-dihydro-3，5-dihydroxy-6-methyl-4（H）-pyran-4-one （P<0.05）. The contents of pungent odor components such as caproic acid， nonanoic acid and synthetic camphor decreased after processing， while the contents of sweet flavor components such as 2-methyl-2-butenal， furfural and 5-hydroxymethylfurfural increased after processing， and the contents of furfural， 5-methyl-2-furanmethanol， 5-hydroxymethylfurfural and other aroma components in the braised products were significantly higher than that in the steamed products. ConclusionHS-GC-MS can quickly identify the volatile substance basis that causes the different odors of PMR and its processed products. The effect of processing methods on the odor is greater than that of origin. There is a significant correlation between the color parameter of L^* and contents of volatile components， the "raw" taste of PMR may be related to volatile components such as caproic acid， pelargonic acid and synthetic camphor， the "flavor" after processing may be related to the increase of the contents of 2-methyl-2-butenal， furfural， 5-hydroxymethylfurfural， methyl maltol and furfuryl alcohol.