Aili Gao

Genome Sequence of the Milbemycin-Producing Bacterium Streptomyces bingchenggensis

Streptomyces bingchenggensis is a soil-dwelling bacterium producing the commercially important anthelmintic macrolide milbemycins. Besides milbemycins, the insecticidal polyether antibiotic nanchangmycin and some other antibiotics have also been isolated from this strain. Here we report the complete genome sequence of S. bingchenggensis. The availability of the genome sequence of S. bingchenggensis should enable us to understand the biosynthesis of these structurally intricate antibiotics better and facilitate rational improvement of this strain to increase their titers.

Reversal of P-glycoprotein-mediated multidrug resistance in vitro by milbemycin compounds in adriamycin-resistant human breast carcinoma (MCF-7/adr) cells

The effects of milbemycin A(4) (MB A(4)), milbemycin oxime A(4) (MBO A(4)) and milbemycin beta(1) (MB beta(1)) on reversing multidrug resistance (MDR) of tumor cells were firstly conducted according to the following research, including MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide] assay, the accumulation of adriamycin, the accumulation and efflux of rhodamine 123 (Rh123), the regulations of MDR1 gene, and expression of P-gp. The three milbemycins (5muM) showed strong potency to increase adriamycin cytotoxicity toward adriamycin-resistant human breast carcinoma cells MCF-7/adr with reversal fold (RF) of 21.42, 19.06 and 14.89, respectively. In addition, the mechanisms of milbemycins on P-glycoprotein (P-gp)-mediated MDR demonstrated that the milbemycins significantly increased the intracellular accumulations of adriamycin and Rh123 via inhibiting P-gp transport function. Based on the analysis of the P-gp and MDR1 gene expression using flow cytometry and RT-PCR, the results revealed that milbemycin compounds, particularly MB A(4), could regulate down the expression of the P-gp and MDR1 gene. These findings suggest that the milbemycins probably represent promising agents for overcoming MDR in cancer therapy, and especially MB A(4) is better modulator with the lowest toxicity.

Reversal of P-glycoprotein-mediated multidrug resistance in vitro by doramectin and nemadectin

Objectives Multidrug resistance (MDR) is a serious obstacle encountered in cancer treatment. This study was performed to explore the reversal of MDR by doramectin from the avermectin family and nemadectin belonging to the milbemycin family.

Reversal effects of two new milbemycin compounds on multidrug resistance in MCF-7/adr cells in vitro.

Development of agents to overcome multidrug resistance (MDR) is important in cancer chemotherapy, and the overexpression of P-glycoprotein (P-gp) is one of the major mechanisms of MDR. In this paper, we evaluated the effects of two new milbemycin compounds, milbemycin β(14) and secomilbemycin D, isolated from fermentation broth of S. bingchenggensis on reversing MDR of adriamycin-resistant human breast carcinoma (MCF-7/adr) cells. We observed that the both milbemycins (5 μM) showed strong potency to increase adriamycin cytotoxicity toward MCF-7/adr cells with reversal fold (RF) of 13.5 and 10.59, respectively. In addition, the mechanisms of milbemycins on reversing P-gp-mediated MDR demonstrated that they significantly increased the accumulations of adriamycin and Rh123 via inhibiting P-gp efflux in MCF-7/adr cells. Furthermore, the results also revealed that milbemycin β(14) and secomilbemycin D could regulate down the expression of P-gp, but not affect the expression of MDR1 gene. In conclusion, our observations suggest that the two new milbemycin compounds probably represent the promising agents for reversing MDR in cancer therapy.

Human amniotic epithelial stem cells inhibit microglia activation through downregulation of tumor necrosis factor-α, interleukin-1β and matrix metalloproteinase-12 in vitro and in a rat model of intracerebral hemorrhage

BACKGROUND AIMS The molecular mechanisms by which stem cell transplantation improves functional recovery after intracerebral hemorrhage (ICH) are not well understood. Accumulating evidence suggests that microglia cells are activated shortly after ICH and that this activation contributes to secondary ICH-induced brain injury. We studied the effect of human amniotic epithelial stem cells (HAESCs) on microglia activation. METHODS To study the effect of HAESCs in vitro, we used thrombin to activate the microglia cells. Twenty-four hours after thrombin treatment, the levels of tumor necrosis factor-α and interleukin-1β were measured by enzyme-linked immunosorbent assay. In vivo, the HAESCs were transplanted into the rat striatum 1 day after collagenase-induced ICH. The expression levels of matrix metalloproteinase (MMP)-12 and microglia infiltration in the peri-hematoma tissues were determined 7 days after ICH through the use of reverse transcriptase-polymerase chain reaction and immunohistochemical analysis, respectively. RESULTS Thrombin-activated microglia expression of tumor necrosis factor-α, interleukin-1β and MMP-12 was significantly reduced through contact-dependent and paracrine mechanisms when the HAESCs were co-cultured with microglia cells. After transplantation of HAESCs in rat brains, the expression levels of MMP-12 and microglia infiltration in the peri-hematoma tissues were significantly reduced. CONCLUSIONS Our observations suggest that microglia activation could be inhibited by HAESCs both in vitro and in vivo, which may be an important mechanism by which the transplantation of HAESCs reduces brain edema and ameliorates the neurologic deficits after ICH. Therefore, we hypothesize that methods for suppressing the activation of microglia and reducing the inflammatory response can be used for designing effective treatment strategies for ICH.

Spinal duraplasty with two novel substitutes restored locomotor function after acute laceration spinal cord injury in rats

A dural tear is a common complication after acute laceration spinal cord injury (ALSCI). An unrepaired dural tear is associated with poor locomotor functional recovery. Spinal duraplasty with biomaterials may promote functional recovery in ALSCI. However, an ideal dural substitute has not yet been found. In this work, we investigated the possibility of using a denuded human amniotic membrane (DHAM) or DHAM seeded on bone marrow stromal cells (DHAM-BMSCs) as duraplasty biomaterials. We patched broken dura with the two novel substitutes in an ALSCI rat model. At the end of the eighth week, we observed that the neural motor function was recovered according to the Basso-Beattie-Bresnahan scale, and the neural loop was successfully reestablished between the ends of the lesions by motor-evoked potentials in the duraplasty groups. Moreover, the DHAM-BMSCs repaired the dura and resulted in a significant reduction in the total lesion and cystic volumes by nearly 10-fold versus the control group (p < 0.01). The levels of neurotrophic factors and NF-200-positive fibers were also improved in the duraplasty groups, compared to the control group. Our data suggest that the two novel substitutes may be promising grafts for patching dural defects to improve locomotor function after ALSCI.

Moxidectin inhibits glioma cell viability by inducing G0/G1 cell cycle arrest and apoptosis

Moxidectin (MOX), a broad-spectrum antiparasitic agent, belongs to the milbemycin family and is similar to avermectins in terms of its chemical structure. Previous research has revealed that milbemycins, including MOX, may potentially function as effective multidrug resistance agents. In the present study, the impact of MOX on the viability of glioma cells was examined by MTT and colony formation assay, and the molecular mechanisms underlying MOX-mediated glioma cell apoptosis were explored by using flow cytometry and apoptosis rates. The results demonstrated that MOX exerts an inhibitory effect on glioma cell viability and colony formations in vitro and xenograft growth in vivo and is not active against normal cells. Additionally, as shown by western blot assay, it was demonstrated that MOX arrests the cell cycle at the G0/G1 phase by downregulating the expression levels of cyclin-dependent kinase (CDK)2, CDK4, CDK6, cyclin D1 and cyclin E. Furthermore, it was revealed that MOX is able to induce cell apoptosis by increasing the Bcl-2-associated × protein/B-cell lymphoma 2 ratio and activating the caspase-3/-9 cascade. In conclusion, these results suggest that MOX may inhibit the viability of glioma cells by inducing cell apoptosis and cell cycle arrest, and may be able to function as a potent and promising agent in the treatment of glioma.

Ivermectin inhibits the growth of glioma cells by inducing cell cycle arrest and apoptosis in vitro and in vivo: SONG et al.

Glioma, the most predominant primary malignant brain tumor, remains uncured due to the absence of effective treatments. Hence, it is imperative to develop successful therapeutic agents. This study aimed to explore the antitumor effects and mechanisms of ivermectin (IVM) in glioma cells in vitro and in vivo. The effects of IVM on cell viability, cell cycle arrest, apoptosis rate, and morphological characteristics were determined respectively by MTT assay/colony formation assay, flow cytometry, and transmission electron microscope. In addition, the expression levels of cycle‐related and apoptosis‐associated proteins were individually examined by Western blot analysis. Moreover, cell proliferation and apoptosis analyses were carried out by TUNEL, Ki‐67, cleaved caspase‐3, and cleaved caspase‐9 immunostaining assay. Our results demonstrated that IVM has a potential dosage‐dependent inhibition effect on the apoptosis rate of glioma cells. Meanwhile, the results also revealed that IVM induced apoptosis by increasing caspase‐3 and caspase‐9 activity, upregulating the expressions of p53 and Bax, downregulating Bcl‐2, activating cleaved caspase‐3 and cleaved caspase‐9, and blocking cell cycle in G0/G1 phase by downregulating levels of CDK2, CDK4, CDK6, cyclin D1, and cyclin E. These findings suggest that IVM has an inhibition effect on the proliferation of glioma cells by triggering cell cycle arrest and inducing cell apoptosis in vitro and in vivo, and probably represents promising agent for treating glioma.