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Reversal of doxorubicin resistance by guggulsterone of Commiphora mukul in vivo.

ABSTRACT

Our previous study has shown co-administration of guggulsterone resulted in significant increase in chemosensitivity of multidrug-resistant human breast cancer MCF-7/DOX cells to doxorubicin (DOX) in vitro. The present study was designed to investigate whether guggulsterone had the similar modulatory activities in vivo. MCF-7/DOX and MCF-7 xenograft mice models were established. At the end of the experiment (day 28), doxorubicin treatment alone did not significantly inhibit tumor growth in MCF-7/DOX xenograft, indicating that it retained doxorubicin resistance. Whereas, doxorubicin treatment alone significantly inhibited tumor growth in MCF-7 xenograft, suggesting that it maintained doxorubicin sensitivity. When doxorubicin and guggulsterone were co-administrated, their antitumor activities were augmented in MCF-7/DOX xenograft. However, combination therapy did not enhance the antitumor effects of doxorubicin in MCF-7 xenograft. The expression of proliferative cell nuclear antigens PCNA and Ki67 after doxorubicin treatment alone was not significantly different from that of vehicle group in MCF-7/DOX xenograft. On the contrary, doxorubicin treatment alone significantly reduced PCNA and Ki67 expression in MCF-7 xenograft. Combination therapy also significantly reduced PCNA and Ki67 expression in MCF-7/DOX xenograft, compared to doxorubicin treatment alone. However, combination therapy did not enhance the inhibitory effects of doxorubicin on PCNA and Ki67 expression in MCF-7 xenograft. Examining the apoptotic index by TUNEL assay showed similar results. Further studies demonstrated the inhibitory effects of guggulsterone on Bcl-2 and P-glycoprotein expression were the possible reason to increase chemosensitivity of MCF-7/DOX cells to doxorubicin in vivo. Examining body weight, hematological parameters, hepatic, cardiac and gastrointestinal tracts histopathology revealed that no significant signs of toxicity were related to guggulsterone. Guggulsterone might reverse doxorubicin resistance in vivo, with no severe side effects.

Keywords:

Guggulsterone

Xenograft

Doxorubicin

Drug-resistance

Introduction

Drug resistance is one of the major problems in cancer chemotherapy. When tumor cells acquire resistance to one chemotherapeutic agent, they usually show cross-resistance to others, the so-called multidrug resistance phenomenon (Ishida et al., 1994). Multidrug resistance can be considered as one of the major reasons for cancer chemotherapy failure. The molecular mechanisms leading to multidrug resistance include the activation of transport and detoxification systems, enhancement of target repair activities, alteration of drug targets, and disregulation of cell death pathways (Ross, 2000). A promising approach to circumvent drug resistance is to utilize non-toxic compounds in combination with the known and well-established chemotherapeutic agents. The limited success in treatment with these chemosensitizers such as verapamil may be attributed to unattainable effective plasma concentrations of these agents due to severe adverse reactions. It was, therefore, warranted to search for additional chemosensitizers with good safety and potential clinical value (O'Connor, 2007).

Guggulsterone [4,17(20)-pregnadiene-3,16-dione], the active component of gugulipid, is derived from the gum resin of the tree Commiphora mukul, and has cis and trans isomers. This gum resin has been used for centuries in Ayurvedic medicine to treat obesity, arthritis, and hyperlipidemia (Mencarelli et al., 2009; Rayalam et al., 2009). Our previous study has shown that guggulsterone could inhibit P-glycoprotein-mediated multidrug resistance in doxorubicin resistant human breast cancer MCF-7/DOX cells. Co-administration of guggulsterone resulted in a significant increase in chemosensitivity of MCF-7/DOX cells to doxorubicin, compared to doxorubicin treatment alone (Xu et al., 2011). However, there is little information regarding the effects of combination therapy for resistant cancer treatment in vivo. Therefore, the present work was designed to investigate the modulatory activities of guggulsterone on the antitumor effects of doxorubicin in resistant breast tumor xenograft model.

Materials and methods

Material

Z-Guggulsterone and doxorubicin as hydrochloride salt were purchased from Sigma-Aldrich Co. (St. Louis, MO, USA). MCF-7 cells were purchased from Shanghai Institute of Cell Biology. MCF-7/DOX cells were developed from the parental MCF-7 cells by stepwise selection for resistance with increasing concentration of doxorubicin and maintained in the presence of doxorubicin (0.2 [micro]g/ml). Cells were grown in RPMI 1640 containing 10% fetal calf serum and 300mg/l glutamine at 37 [degrees]C in a humidified C[O.sub.2] incubator.

Athymic nude mice were used for MCF-7/DOX and MCF-7 xenografts. BALB/c (nu + nu + genotype) mice were obtained from Shanghai Laboratory Animal Center (Shanghai, China). Animals were maintained in pathogen free environment at the animal care facility, where they were provided with sterilized food and water. They were maintained under a 12 h light-dark cycle. Female mice of 4-6-weeks-old, weighing 20-25 g, were used. All animal protocols were approved by the local Ethics Committee of Shanghai Tenth People's Hospital (Tongji University).

Reversal of doxorubicin resistance assay

MCF-7/DOX and MCF-7 xenograft models were established according to published protocols (Ulmann et al., 1991). In detail, transplantable cells (1 x 107) were collected and subcutaneously injected in the right flanks of mice. When tumor volume reached 100 [mm.sup.3] in size approximately, mice were randomized into five different treatment groups, each of eight mice. These are Group a: control group (mice without treatment), Group b: vehicle group (treated with DMSO only), Group c: doxorubicin alone (5 mg/kg), Group d: guggulsterone alone (30 mg/kg), and their combination in Group e. Treatment was given intraperitoneally twice a week for 2 weeks. The selected doses were based on our pilot study as well as previous report (Xu et al., 2011; An et al., 2009). For all mice, tumors were measured in two dimensions using calipers every four days and tumor volumes were calculated using the following formula: volume = a x [b.sup.2]/2, where a is the width at the widest point of the tumor and b is the width perpendicular to a. The curve of tumor growth was drawn according to the percentage of change in tumor volume from first day of treatment (day 1) in different treatment groups.

Histological examination and TUNEL assay of apoptosis

Animals were sacrificed and tumor tissues from each mouse were excised at day 28, then tumors were fixed with formalin and embedded in paraffin. Serial sections were cut from each paraffin block and stained with hematoxylin-eosin (H&E) staining for histopathology, and fluorescent TUNEL staining for apoptosis (Roche). Microphotographs were obtained using a Zeiss LSM 510 confocal microscope. Apoptosis index was determined by counting the number of apoptotic cells compared to the number of normal viable ones in at least five high power fields (20x) of each section in a blinded fashion. Three tumors per group were analyzed to take their average.

Immunohistochemistry for anti-proliferative activity and Bcl-2

Proliferative cell nuclear antigens PCNA and Ki67 were used to assess the anti-proliferative activities in different treatment groups. Bcl-2 protein expression in tumor sections was examined to analyze the mechanism by which guggulsterone inhibits tumor growth. Briefly, tumor sections were incubated overnight at 4[degrees]C with mouse antihuman PCNA (1:6000 dilution; Santa Cruz Biotechnology), Ki67 (1:200 dilution; Santa Cruz Biotechnology) or Bcl-2 (1:100 dilution; Santa Cruz Biotechnology) antibodies. immunoreactions were visualized using a streptavidin-biotin complex method followed by diaminobenzidine reaction. Tumor sections were counterstained with hematoxylin to visualize the nuclei. Immunoreactions were determined by counting the number of positive antigens to total cell number in five high power fields (40x) of each section in a blinded fashion. Three tumors per group were analyzed to take their average.

Western blot analysis of Bcl-2 and P-glycoprotein levels

Three hundred milligram of tumor tissue was homogenized in ice-cold phosphate-buffered saline, and then centrifuged at 1000rpm for 5 min at room temperature. Each pellet was resuspended with 1 ml lysis buffer (20 mM Tris-HCl, pH 7.5, 1 mM sodium orthovanadate, 50 mM NaF, 20 mM leupeptin, 10 mM ([beta]-mercaptoethanol, 2.5 mM sodium pyrophosphate, 0.2 mM phenylmethanesulfonyl fluoride) plus 1% Triton-100, and then was incubated at 0[degrees]C for 30 min. Western blot analysis was carried out as previously described (Brigui et al., 2003). Briefly, the lysates were centrifuged at 10,000 x g for 10 min at 4[degrees]C and the supernatants were checked for protein content. Equal amounts of proteins (50 mg) were separated using a 7.5% SDS-polyacrylamide gel/5% stacking gel. The resolved proteins were transferred to a nitrocellulose membrane from the gel, and immunoblotted with antibodies for Bcl-2 (ab32124, Abeam, China) and P-glycoprotein (ab3083, Abeam, China). Signals were detected by chemiluminescence (ECL kit; Abeam).

Doxorubicin accumulation assay

To measure intracellular doxorubicin accumulation in MCF-7/DOX xenograft tumor cells, the tumors were resected, and MCF-7/DOX cells were collected. A total of 1 x [10.sup.8] cells per sample were washed 3 times with cold phosphate-buffered saline, resuspended in 0.3 N HC1 and 50% ethanol, and disrupted by sonication for 30 s. After centrifugation at 10,000 rpm for 15 min, the doxorubicin concentration in the supernatant was measured by fluorospectrophotometry (Synergy 4, BioTek Ltd., USA) with excitation and emission wavelengths of 500 and 590 nm, respectively. The doxorubicin concentration (p.g/108 cells) was calculated from a standard curve prepared using known amounts of doxorubicin. Three tumors per group were analyzed to take their average.

Toxicity evaluation

Mice body weight was measured every four days to evaluate the systemic treatment toxicity. Other signs of unwanted toxicity were monitored including fur-roughing, shedding and local trauma at the site of injection. Decrease in general animal activity was also examined as general sign of systemic toxicity. Blood samples were collected from the retro-orbital plexus of individual animals and analyzed for hematological parameters. After complete gross necropsy examination, the hearts, livers and gastrointestinal tracts in all treatment groups were preserved in 10% formalin. Tissue samples were taken for H&E staining to allow histological evaluation of possible cardiotoxicity, hepatotoxicity and mucositis.

Statistical analysis

Data were expressed in the form of mean [+ or -] S.D., and the significance of the difference between groups was determined by ANOVA followed by the Bonferroni post hoc test.

Results

Effectiveness and toxicity evaluation

The effects of doxorubicin and/or guggulsterone treatment on tumor growth are presented in Fig. 1 a and b. At the end of the experiment (day 28), doxorubicin treatment alone did not significantly inhibit tumor growth in MCF-7/DOX xenograft, indicating that it retained doxorubicin resistance (Fig. la). The change of tumor volume after doxorubicin treatment alone (385.0 [+ or -] 58.50%) was not significantly different from that of vehicle group (422.9 [+ or -] 31.50%). However, doxorubicin treatment alone significantly inhibited tumor growth in MCF-7 xenograft, suggesting that it maintained doxorubicin sensitivity, and the change of tumor volume was reduced to 84.37 [+ or -] 19.53% (Fig. lb). Mice receiving guggulsterone alone demonstrated almost no change of tumor volume in MCF-7/DOX and MCF-7 xenografts, where reached 405.4 [+ or -] 23.63% and 408.6 [+ or -] 27.14%, respectively (Fig. la and b). When doxorubicin and guggulsterone were co-administrated, their antitumor activities were augmented in MCF-7/DOX xenograft. The change of tumor volume was reduced to 93.87 [+ or -] 20.86% (Fig. la). Whereas, combination therapy did not enhance the antitumor effects of doxorubicin in MCF-7 xenograft, and the change of tumor volume reached 86.41 [+ or -] 27.04% (Fig. 1b).

Fig. 1c and d presents the change of mice body weight in MCF-7/DOX and MCF-7 xenografts. Treatment with doxorubicin and/or guggulsterone resulted in no significant differences in body weight, and caused no death. No mice manifested signs of other adverse effects (inactivity, fur-roughing or tissue damage at injection site). Doxorubicin at the dose of 5 mg/kg caused a significant reduction in total white blood cells and platelets count. Combination therapy resulted in a restoration of total white blood cells and platelets count, compared to doxorubicin treatment alone (Table 1). No substantial abnormalities were recognized on examining heart, liver and gastrointestinal tract sections in different treatment groups (data not shown).

Histological examination of tumor tissues

Tumors in vehicle group from MCF-7/DOX and MCF-7 xenografts showed an area of necrosis, and interspersed with many regions of viable tumor cells near blood vessels and a margin of actively proliferating cells at the tumor periphery (Fig. 2b). Tumors in control and guggulsterone group showed no observed changes in the pattern of tumor cells, compared to vehicle group (Fig. 2a and d). Small area of tumor cells in doxorubicin group from MCF-7/DOX xenograft showed cytoplasm vacuolization and nuclei degeneration. The nuclei shrunk in size and became pyknotic (Fig. 2c). However, Tumors in doxorubicin group from MCF-7 xenograft showed complete degeneration of tumor cells in the subcutaneous region with remnant of pyknotic nuclei. Tumors in combination group from MCF-7/DOX and MCF-7 xenografts showed the same results (Fig. 2e).

Immunohistochemistry of proliferative markers

To determine the anti-proliferative effects of doxorubicin and/or guggulsterone treatment, immunohistochemical staining with an antibody against PCNA was performed. Most tumor tissues from MCF-7/DOX and MCF-7 xenografts stained positive for PCNA antibody (Fig. 3a). Meanwhile, guggulsterone treatment alone did not significantly reduce PCNA expression (Fig. 3d) in tumor tissues from MCF-7/DOX and MCF-7 xenografts, compared to vehicle group (Fig. 3b). The proliferative index for doxorubicin group from MCF7/DOX xenograft (0.76 [+ or -] 0.08, Fig. 3c) was not significantly different from that of vehicle group (0.83 [+ or -] 0.06, Fig. 3b). However, the proliferative index for doxorubicin group from MCF-7 xenograft (0.40 [+ or -] 0.04, Fig. 3c) was significantly different from that of vehicle group (0.89 [+ or -] 0.13, Fig. 3b). Furthermore, combination group from MCF-7/DOX and MCF-7 xenografts also reduced PCNA expression (Fig. 3e), which was significantly different from that of doxorubicin group. Fig. 3f represents the proliferative indices of PCNA in different treatment groups.

Examining Ki67 expression showed the similar results. Guggulsterone treatment alone did not significantly reduce Ki67 expression (Fig. 4d) from MCF-7/DOX and MCF-7 xenografts, compared to vehicle group (Fig. 3b). The proliferative index for doxorubicin group from MCF-7/DOX xenograft (0.69 [+ or -] 0.09, Fig. 4c) was not significantly different from that of vehicle group (0.77 [+ or -] 0.10, Fig. 4b). However, the proliferative index for doxorubicin group from MCF-7 xenograft (0.39 [+ or -] 0.07, Fig. 4c) was significant different from that of vehicle group (0.82 [+ or -] 0.14, Fig. 4b). Combination group from MCF-7/DOX and MCF-7 xenografts also reduced Ki67 expression (Fig. 4e), which was significantly different from that of doxorubicin group. Fig. 4f represents the proliferative indices of Ki67 in different treatment groups.

TUNEL assay of apoptosis

The number of apoptotic cells in tumor sections was determined by TUNEL assay. The apoptotic indices in guggulsterone group (Fig. 5d) from MCF-7/DOX and MCF-7 xenografts were not significantly different from those of vehicle group (Fig. 5b). Meanwhile, doxorubicin group from MCF-7/DOX xenograft induced an apoptotic index of 0.09 [+ or -] 0.03 (Fig. 5c), which was also not significantly different from vehicle group (0.07 [+ or -] 0.02; Fig. 5b). However, the apoptotic index in doxorubin group (0.49 [+ or -] 0.16, Fig. 5c) from MCF-7 xenograft was significantly different from that of vehicle group (0.05 [+ or -] 0.03, Fig. 5b). Furthermore, combination group from MCF-7/DOX and MCF-7 xenografts increased the apoptotic index to 0.46 [+ or -] 0.07 and 0.51 [+ or -] 0.11, respectively (Fig. 5e), which was significantly different from that of doxorubicin alone. Fig. 5f represents the apoptotic indices in different treatment groups.

Bcl-2 expression assay

To determine the mechanism by which guggulsterone inhibits tumor growth, we further assessed Bcl-2 protein expression in tumor sections from MCF-7/DOX xenograft. As expected, Bcl-2 was expressed in tumor cells cytoplasm, which is known to play an inhibitory role in apoptosis. Guggulsterone treatment alone did not significantly decrease Bcl-2 expression (0.54 [+ or -] 0.10, Fig. 6d), compared to vehicle group (0.58 [+ or -]0.08, Fig. 6b). However, combination therapy down-regulated Bcl-2 expression (0.31 [+ or -]0.09, Fig. 6e), which was significantly different from doxorubicin treatment alone (0.51 [+ or -] 0.08, Fig. 6c). Fig. 6f represents Bcl-2 expression in different treatment groups. The results of Western blot analysis similarly showed that combination therapy inhibited Bcl-2 expression (Fig. 7).

P-glycoprotein expression analysis

To determine whether P-glycoprotein is present in MCF-7/DOX cells derived from tumors and effects of guggulsterone on P-glycoprotein expression, Western blot analysis was carried out. As seen in Fig. 7, P-glycoprotein level in tumor cells from MCF-7/DOX xenograft was significant higher than that from MCF-7 xenograft. After treated with doxorubicin, P-glycoprotein level in tumor cells from MCF-7/DOX xenograft was not different from that of vehicle group (Fig. 8). However, obvious change of P-glycoprotein level was observed in tumor cells from MCF-7/DOX xenograft treated with guggulsterone alone or combination therapy, compared to doxorubicin alone, which suggests that guggulsterone inhibits P-glycoprotein expression (Fig. 8).

Doxorubicin accumulation analysis

Doxorubicin is a good substance for P-glycoprotein, and agents that inhibit P-glycoprotein function have been found to increase accumulation of doxorubicin in drug-resistance cells. Therefore, we finally investigated whether guggulsterone enhanced doxorubicin antitumor activities in MCF-7/DOX xenograft by increasing intracellular accumulation of doxorubicin. The results showed that doxorubicin concentrations in tumors treated with doxorubicin (5 mg/kg), guggulsterone (30 mg/kg), and their combination were 1.427 [+ or -] 0.189, 1.023 [+ or -] 0.085, and 2.953 [+ or -] 0.171 [micro]g/g tumor tissue, respectively. Combination therapy resulted in 2.1-fold increase in intracellular doxorubicin accumulation, compared to that of doxorubicin treatment alone (Fig. 9).

Discussion

Clinically, multidrug resistance is one of the major obstacles for chemotherapy. Therefore, searching for effective and applicably drug-resistant modulators has been given the highest priority for enhancing the sensitivity of drug-resistant tumor cells to chemotherapeutics. Unfortunately, clinical trials with these compounds could lead to unacceptable side effects or toxicity (Kerb et al., 2001; Ling, 1997).

Guggulsterone has several advantages as a potential drug-resistant modulator. Its anti-proliferative dosages show almost no toxicity in vitro and in vivo (Xu et al., 2009; An et al., 2009; Xu et al., 2011). Doxorubicin is one of the most valuable chemotherapeutic drugs in breast cancer. Unfortunately, its progress has the price of development of tumor resistance impairing successful therapy. Therefore, our present work was designed to investigate the modulatory effects of guggulsterone on the antitumor effects of doxorubicin in MCF-7/DOX and MCF-7 xenografts. Our findings demonstrated that doxorubicin treatment alone did not significantly inhibit MCF-7/DOX tumor growth, as predicted from its resistance phenotype. Our results are in agreement with Mimnaugh et al. who reported that doxorubicin failed to slow doxorubicin resistant MCF-7 xenograft growth (Mimnaugh et al., 1991).

Furthermore, in the present study, guggulsterone treatment alone also did not inhibit tumor growth. However, when doxorubicin and guggulsterone were co-administrated, their growth inhibiting activity was significantly augmented in MCF-7/DOX xenograft. Our work is supported by study showing that guggulsterone inhibits xenograft growth derived from pancreatic cancer cell lines (Ahn et al., 2012). The enhanced anti-proliferative effect of combined treatment was further substantiated by assessing PCNA and Ki67 expression. The proliferative indices for doxorubicin treatment alone were not significantly different from that of vehicle group in MCF-7/DOX xenograft. This confirms the observed ineffectiveness of doxorubicin against the studied tumor model. However, combination therapy resulted in significant reduction of both markers in MCF-7/DOX xenograft. Similarly, TUNEL assay indicated that this combination significantly increased the apoptotic index, compared to doxorubicin treatment alone in MCF-7/DOX xenograft. These observations are consistent with our previous in vitro study (Xu et al., 2011). Moreover, the present study shows that combination-therapy-mediated suppression of MCF-7/DOX xenograft growth is accompanied by Bcl-2 expression inhibition. There is evidence that over-expression of the anti-apoptotic protein Bcl-2 might be associated with drug resistance in some human cancer cell lines (Minn et al., 1995; Miyashita et al., 1994; Sumantran et al., 1995). The results suggest that the apoptotic effect of combination treatment toward MCF-7/DOX cells is, at least partly, caused by an alternation in the expression of Bcl-2 protein.

In order to obtain insight in the possible mechanism of guggulsterone enhancing the cytotoxicity of doxorubicin, P-glycoprotein expression and function were further measured in the present study. The results indicated that P-glycoprotein level in tumor cells from MCF-7/DOX xenograft was higher than that from MCF-7 xenograft, and was down-regulated after guggulsterone treatment. Furthermore, combination therapy increased doxorubicin concentration in tumor cells from MCF-7/DOX xenograft. These findings are in agreement with our previous in vitro study demonstrating that guggulsterone could inhibit P-glycoprotein expression and function in MCF-7/DOX cells (Xu et al., 2011). This suggests that P-glycoprotein inhibition might be the possible mechanism by which guggulsterone sensitized the resistant tumors to doxorubicin in MCF-7/DOX xenograft model. However, because various potential targets have been identified for the antitumor and chemopreventive activities of guggulsterone, the molecular mechanism will require further study.

Based on our findings, it is possible to conclude that apoptotic induction is an essential event in the guggulsterone-mediated suppression of MCF-7/DOX cell growth in vitro and in vivo. However, our observations should be confirmed in further studies to more definitely establish the in vivo relevance of the in vitro findings. Moreover, we used relatively high guggulsterone doses in treating mice. If guggulsterone could be administered intravenously or into tumor feeding vessels, higher concentrations could be achieved with lower doses. Further dosing studies are also needed if we are to extrapolate our findings to humans.

To further characterize the effects of combination therapy, toxicity profile was examined. No significant differences were found in body weight in different treatment groups. Likewise, no general signs of toxicity or deaths were observed in any groups. Several concerns were raised regarding the dose limiting cardiotoxicity of doxorubicin (Von Hoff et al., 1982). Histopathological examination of heart sections from different treatment groups showed no observed signs of toxicity. Our findings might appear contradictory to the reported cardiotoxicity of doxorubicin (Yi et al., 2006). However, this can be explained on the basis of less cumulative dose and different dosing schedule of doxorubicin used in the current study. Furthermore, examining liver and gastrointestinal sections did not indicate any observable histopathological changes in the present work. In addition, combination therapy ameliorated total white blood cells and platelets count, compared to doxorubicin treatment alone. The promising anti-tumor activities and little toxicity suggest that guggulsterone has the potential to be a novel adjuvant therapeutic agent in breast cancer chemotherapy.

In conclusion, we demonstrate for the first time that guggulsterone reverses doxorubicin resistance in human breast tumor xenograft models. This is evidenced by synergizing its antiproliferative and apoptotic effects without significantly enhancing its toxicity. Although the mechanism of guggulsterone inhibiting drug resistance is not clear, our present study suggests that it is possible to improve efficacy of some chemotherapeutic drugs on cancer by co-administration of guggulsterone.

http://dx.doi.org/10.1016/j.phymed.2014.06.003

ARTICLE INFO

Article history:

Received 21 November 2013

Received in revised form 6 May 2014

Accepted 9 June 2014

Acknowledgement

This work was supported by National Natural Science Foundation of China (No. 81073104).

References

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Hong-Bin Xu (a), * (1), Zhao-Li Shen (b), (1), Jin Fu (c), (1), Lu-Zhong Xu (a)

(a) Department of Pharmacy, Shanghai Tenth People's Hospital, Tongji University School of Medicine, Shanghai 200072, China

(b) Department of Neurosurgery, Shanghai Tenth People's Hospital, Tongji University School of Medicine, Shanghai 200072, China

(c) Department of Neurology, Shanghai Tenth People's Hospital, Tongji University School of Medicine, Shanghai 200072, China

* Corresponding author. Tel.: +86 2166302761; fax: +86 2166307668.

E-mail addresses: xuhongbin@tongji.edu.cn, xuhongbin119@yahoo.cn (H.-B. Xu).

(1) These authors have equally contributed to this article.

Table 1
Effects of guggulsterone on doxorubicin-induced
myelosuppression in mice.

                                        MCF-7

                         RBCs count             WBCs count
                          (million/             (thousand/
Treatment group           [micro]l)             [micro]l)

Control              6.12 [+ or -] 0.309   10.86 [+ or -] 0.248
Vehicle              5.91 [+ or -] 0.185   10.02 [+ or -] 0.249
Doxorubicin
  (30 mg/kg)
Guggulsterone        5.82 [+ or -] 0.226   7.29 [+ or -] 0.3541
  (5 mg/kg)          6.13 [+ or -] 0.193   10.39 [+ or -] 0.402
Doxorubicin
  (5 mg/kg)
  + guggulsterone
  (30 mg/kg)         6.07 [+ or -] 0.337   9.94 [+ or -] 0.281b

                             MCF-7                MCF-7/DCX

                        Platelets count          RBCs count
                          (thousand/             (million /
Treatment group            [micro]l)              [micro]l)

Control              513.53 [+ or -] 11.42   5.52 [+ or -] 0.413
Vehicle              528.16 [+ or -] 19.63   5.38 [+ or -] 0.264
Doxorubicin
  (30 mg/kg)
Guggulsterone        401.73 [+ or -] 9.841   4.86 [+ or -] 0.097
  (5 mg/kg)          531.84 [+ or -] 23.19   5.73 [+ or -] 0.219
Doxorubicin
  (5 mg/kg)
  + guggulsterone
  (30 mg/kg)         493.82 [+ or -] 6.31b   5.06 [+ or -] 0.362

                                         MCF-7/DCX

                          WBCs count           Platelets count
                          (thousand/              (thousand/
Treatment group           [micro]l)               [micro]l)

Control              11.89 [+ or -] 0.507   462.29 [+ or -] 18.86
Vehicle              11.52 [+ or -] 0.356   451.22 [+ or -] 10.26
Doxorubicin
  (30 mg/kg)
Guggulsterone        7.64 [+ or -] 0.4281   324.81 [+ or -] 13.321
  (5 mg/kg)          11.93 [+ or -] 0.119   473.91 [+ or -] 15.28
Doxorubicin
  (5 mg/kg)
  + guggulsterone
  (30 mg/kg)         10.53 [+ or -] 0332b   460.62 [+ or -] 9.926

RBCs, red blood cells; WBCs, white blood cells.

(a) Significantly different from vehicle group at p < 0.05.

(b) Significantly different from doxorubingroup at p < 0.05.
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Author:Xu, Hong-Bin; Shen, Zhao-Li; Fu, Jin; Xu, Lu-Zhong
Publication:Phytomedicine: International Journal of Phytotherapy & Phytopharmacology
Article Type:Report
Date:Sep 25, 2014
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