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Curcumin and Oxidative Stress

Curcumin and Oxidative Stress

Article Anc Google Mind-body connection for mood enhancement Demedts M, Sports nutrition for intolerant athletes J, Buhl R, Costabel U, Ocidative R, Jansen HM, Odidative al. Curcumin potentiates laryngeal Curcumin and Oxidative Stress carcinoma radiosensitivity via Oxieative inhibition by suppressing IKKγ expression. We observe significant decreases in the expression of myofibroblast activation and proliferation associated genes in both the NHLF and IPF-F Fig. In terms of autophagy, curcumin has been reported to induce elevated ROS levels and the appearance of autophagy markers and autophagosomes, causing tumor cells to undergo autophagy.


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Curcumin and Oxidative Stress -

Similar results were obtained Fig. We found that curcumin treatment resulted in enhanced accumulation of H 2 O 2 in culture supernatant, peaked at 2 hours Fig. Curcumin generates ROS in HuT and MyLa cells and induces cell death.

A, chemical structure of curcumin. Data represent the results from 1 of the 3 similar experiments. C, at similar time points as in B, H 2 O 2 accumulation in supernatants was measured using Amplex Red Hydrogen Peroxide Assay Kit. Data represent mean values ± SD of 3 similar experiments. Bar graph represents killing of HuT and MyLa cells.

Values are expressed as mean ± SD of 3 similar experiments. E, HuT cells were treated similarly for 24 hours, and apoptosis was detected by TUNEL assay through flow cytometry at FL-1 channel.

The M1 and M2 gates demarcate apoptotic and nonapoptotic populations, respectively. Recently, it has been reported that curcumin is cytotoxic for CTCL cells So, we were interested to see whether ROS is involved in curcumin-mediated killing of HuT and MyLa cells.

Concentration-dependent cytotoxic effects were observed in curcumin-treated both cell lines Fig. Curcumin-mediated cell death was found to be ROS-dependent, because the addition of NAC completely suppressed the cytotoxic effect of curcumin.

Terminal deoxynucleotidyl transferase—mediated dUTP nick end labeling TUNEL assay of curcumin-treated cells further confirmed that curcumin-mediated killing of HuT cells is ROS-dependent. As shown in Fig.

To determine the involvement of caspases in curcumin-mediated apoptosis of HuT cells, we examined the activation of different caspases. Caspase-8 activation was blocked when NAC or broad-range caspase inhibitor, z-VAD-fmk, was present in the system. When the activation of another initiator caspase, caspase-2, was measured by immunoblotting, we obtained dose-dependent activation of caspase-2 in HuT cells after hour treatment with curcumin.

Prior treatment of cells either with NAC or different caspase inhibitors z-VAD-fmk, z-IETD-fmk, and Ac-DEVD-CHO blocked this activation Fig. The cleavage of Bcl-2 family member Bid to t-Bid, a substrate for caspase-8, is involved in linking extrinsic pathway to mitochondrial pathway 6.

Therefore, we investigated the cleavage of Bid to t-Bid in HuT cells treated with curcumin. The cleavage of Bid was suppressed in the presence of NAC or z-VAD-fmk Fig. Curcumin induces caspase activation and alters MMP in a ROS-dependent manner. OD, optical density.

C, cells were treated with curcumin for 24 hours and Bid cleavage was examined by immunoblotting. The MMP changes were determined by DiOC 6 staining using flow cytometry at FL-1 channel. E, cytosolic fraction was used for analysis of cytochrome cyt c by immunoblotting.

Next, we wanted to see whether mitochondrial pathway is involved in curcumin-mediated apoptosis of HuT cells. Alteration of MMP is known to cause release of cytochrome c into the cytosol.

Cytochrome c release from mitochondria into the cytosol of curcumin-treated HuT cells was detected by immunoblotting. Pretreatment with NAC blocked the cytochrome c release, but z-VAD-fmk pretreatment could not significantly suppress the release of cytochrome c Fig.

Next, we measured the activation of caspase-9 and then the activation of caspase-3 in curcumin-treated HuT cells after 24 hours. Concentration-dependent activation of both caspase-9 and -3 was observed Fig. Activation of caspase-3 was markedly reduced in the presence of NAC or broad-range caspase inhibitor, z-VAD-fmk Fig.

PARP cleavage was detected by immunoblotting using anti-PARP antibody. PARP cleavage could be detected in cells treated with Remarkably, this curcumin-mediated cleavage of PARP was completely reverted in the presence of NAC Fig. Curcumin causes caspase-9 and -3 activation and PARP cleavage.

C, PARP cleavage was detected by Western blotting. Apoptosis was determined by Annexin V staining through flow cytometry. The M1 and M2 gates demarcate nonapoptotic and apoptotic populations, respectively.

Data show mean values ± SD of 3 similar experiments. We found that curcumin activated initiator caspase-8, -2, and -9 and executioner caspase, caspase-3, so the next obvious question we asked was that whether caspase-mediated cell death pathway is solely responsible for killing of HuT cells induced by curcumin.

For this purpose, the effect of different caspase inhibitors on curcumin-mediated death of HuT cells was studied. After 24 hours, Annexin V staining was conducted to confirm cell death. Western blot analysis revealed that curcumin effectively downregulated the expression of both c-FLIP L and c-FLIP S as compared with control Fig.

Curcumin also inhibited Bcl-xL expression in a dose-dependent manner Fig. The expression of c-FLIP, Bcl-xL, and cIAP-2 appeared to be ROS-dependent, because the addition of NAC suppressed the inhibitory effect of curcumin on these proteins.

We also checked the expression of XIAP in cell lysates prepared from curcumin-treated HuT cells after 24 hours. The cleaved fragment of XIAP was not visible in NAC-pretreated cells Fig.

XIAP cleavage was completely blocked in the presence of z-VAD-fmk, a broad-range caspase inhibitor, whereas caspase-8—specific inhibitor, z-IETD-fmk, slightly inhibited the XIAP cleavage. In contrast, caspase-3—specific inhibitor, Ac-DEVD-CHO, was not able to block the XIAP cleavage Fig.

Curcumin-mediated ROS generation downregulates antiapoptotic proteins. and immunoblotting was conducted. The arrow indicates full-length and cleaved fragment of XIAP. F, similar experiment as in D was carried out to see the effect of curcumin on Hsp90 and Hsp70 expression by Western blotting.

Arrow indicates full-length and cleaved Hsp It is shown that Hsp90 promotes survival of cancer cells providing stability to pro- and antiapoptotic proteins We, therefore, studied the effect of curcumin on Hsp90 expression.

This curcumin-mediated cleavage of Hsp90 was blocked by antioxidant, NAC Fig. To see whether curcumin-generated ROS specifically cleaves Hsp90, we also checked the expression of Hsp70 in curcumin-treated HuT cells.

Hsp90 cleavage was inhibited by the addition of NAC or broad-range caspase inhibitor, z-VAD-fmk, with curcumin Fig. It has been reported that disruption of Hsp90 leads to proteolytic cleavage of its client proteins Thus, we checked the stability of Hsp90 client proteins IKK-α and IKK-β in curcumin-treated HuT cells.

Immunoblot analysis clearly showed that curcumin downregulated the expression of IKK-α and IKK-β in ROS-dependent manner as pretreatment of cells with NAC abolished the effect of curcumin Fig.

C, HuT cells were treated with varying concentrations of Hsp90 inhibitor, AAG for 12 hours, and NF-κB activation was seen in nuclear extracts by EMSA.

FP, free probe alone no nuclear extracts. It is reported that curcumin inhibits constitutive NF-κB in CTCL cells We speculated that inhibition of NF-κB activity may be due to IKK degradation in curcumin-treated cells.

To see the status of NF-κB, nuclear extracts were prepared from various concentrations of curcumin-treated HuT cells and subjected to EMSA. Curcumin-mediated inhibition of constitutive NF-κB was found to be under the control of ROS, as NAC pretreatment suppressed the inhibitory effect of curcumin.

The specificity of the binding was examined by competition with unlabeled oligonucleotide data not shown. Next, we were interested to see whether Hsp90 is directly involved in NF-κB downregulation.

For this, HuT cells were treated with different concentrations of Hsp90 inhibitor, AAG and nuclear extracts were prepared and analyzed for DNA-binding activity of NF-κB by EMSA. Interestingly, at 2. Recently, it has been shown that Hsp90 forms a complex with beclin-1, a key protein involved in autophagy, and thus maintains the stability of beclin-1 Because curcumin selectively cleaves Hsp90, therefore, we were interested to see the stability of beclin-1 in curcumin-treated HuT cells.

We observed that curcumin treatment resulted in the degradation of beclin-1, which was inhibited by NAC Fig. Time kinetics experiments clearly indicated that degradation of beclin-1 occurred not before 24 hours of curcumin treatment Fig.

Similar result was obtained when similar experiment was carried out in MyLa cells Fig. Curcumin-mediated oxidative stress degrades beclin-1, accumulates LC3-I, and inhibits Atg7. Immunoblot analysis with antibodies against LC3 or actin was conducted.

The ratios of intensities of LC3-II and actin are indicated below each lane. Beclin-1 plays pivotal role in autophagy formation Downregulation of beclin-1 in curcumin-treated cells indicated that the autophagy formation may be inhibited in this cell.

Conversion of autophagy-specific marker LC3-I to LC3-II is an indicator for autophagy formation. Accumulation of LC3-I was observed when higher concentration of curcumin Rapamycin, a known inducer of autophagy was taken as positive control to induce autophagy and to check the conversion of LC3-I to LC3-II in HuT cells.

As expected, rapamycin treatment induced conversion of LC3-I to LC3-II, and this conversion was inhibited in the presence of autophagy inhibitors, 3-MA and wortmannin, Wm Fig.

Next, we studied the expression of autophagy-related proteins Atg5 and Atg7. Curcumin treatment inhibited the expression of Atg7 but not Atg5. Curcumin failed to show its effect in the presence of NAC Fig.

Recent years have seen a growing trend in developing therapeutic agents from natural sources against various diseases. Curcumin, a yellow pigment from turmeric that shows a wide spectrum of biologic function, has proved to be a promising candidate as an anticancer agent.

Curcumin induces cell death in different forms of human cancer cells. Accumulating experimental data indicate that curcumin exerts its cytotoxic effect either by acting as an antioxidant or as a pro-oxidant In this study, we have investigated whether curcumin treatment generates oxidative stress in HuT cells, and if so, then what are the important cell survival mechanisms affected by curcumin-generated oxidative stress?

We report here that curcumin through ROS-dependent mechanism perturbs multiple cell signaling molecules and eventually induces apoptosis in HuT cells. This conclusion is based on several crucial observations. First, time-dependent accumulation of ROS was observed in curcumin-treated cells and secondly, ROS scavenger, NAC, extensively attenuated all curcumin-induced effects such as activation of caspases, downregulation of antiapoptotic gene expressions, cleavage of Hsp90, inhibition of DNA-binding activity of NF-κB, including apoptosis in HuT cells.

We have found that HuT cells are much more vulnerable to oxidative stress than normal peripheral blood mononuclear cell PBMC; data not shown. Induction of substantial apoptosis by curcumin in CTCL cells but not in PBMC from healthy donors, as reported recently by Zhang and colleagues, seems to be due to the generation of excessive ROS in those cancer cells Apoptosis is mediated by the activation of different effector caspases.

According to their function and mode of activation, caspases are classified as initiator caspases that include caspase-2, -8, -9, and executioner caspases such as caspase-3, -6, and Active caspases are generated in 2 distinct pathways, the intrinsic or mitochondrial death pathway and the extrinsic or receptor-mediated pathway 6, We have found that curcumin through ROS generation activated caspase-8, -2, and -9 as well as caspase Caspase-8 activation is a crucial step for the initiation of the extrinsic pathway; on the other hand, caspase-9 activation is essential for the execution of intrinsic pathway.

The link between these 2 pathways is mediated by the truncated proapoptotic protein Bid 6. Caspase-2 like caspase-8 is also known to cleave Bid to t-Bid that alters MMP We observed truncation of Bid, mitochondrial hyperpolarization, cytochrome c release, caspase-9 activation followed by caspase-3 activation, and ultimately cleavage of PARP in curcumin-treated HuT cells.

All these mechanisms are controlled by ROS, as prior treatment with NAC completely blocked this cascade of events. Many researches have shown that curcumin influences multiple signaling pathways to exert its antiapoptotic effect on various cell types Because we found that the caspase cascade is minimally involved in curcumin-mediated killing of HuT cells; therefore, next we investigated the effect of curcumin on other cell survival molecules to find an explanation for curcumin-mediated killing of HuT cells.

It is known that HuT cells express high amount of antiapoptotic proteins such as c-FLIP, Bcl-xL, cIAP, and XIAP. Bcl-xL, a member of Bcl2 family protein, is known to promote cell survival and regulate MMP We found that curcumin at high concentration reduces the expression of Bcl-xL, which probably accounts for the alteration of mitochondrial membrane polarization seen in curcumin-treated cells.

XIAP is known to regulate intracellular ROS by upregulating the expression of antioxidative genes Many cancer cells have been shown to produce high amount of XIAP We observed the cleavage of XIAP in curcumin-treated HuT cells, which probably accounts for the high level of ROS generation by curcumin in these cells.

Researches have shown that Hsp90 function is required for the activation of constitutive IKK, which then activates constitutive NF-κB We found that curcumin treatment results in degradation of IKK and as a consequence, downregulation of NF-κB takes place in HuT cells in a ROS-dependent manner.

Earlier Zhang and colleagues reported that curcumin inhibits constitutive NF-κB in CTCL cells 15 ; but for the first time, we showed the involvement of curcumin-generated ROS in the downregulation of constitutive NF-κB in HuT cells. Curcumin has been shown to induce autophagy in different cell types 29, It is also known that the IKK complex contributes to the induction of autophagy Moreover, like IKK, beclin-1, the key autophagy-promoting protein, has been identified recently as a client protein of Hsp90 Time kinetics show that noteworthy downregulation of beclin-1 occurs at hour time point, at the same time, when Hsp90 function is inhibited by curcumin in a ROS-dependent mechanism.

Next, we investigated autophagy formation in curcumin-treated HuT cells by following the conversion of autophagy-specific marker LC3-I to LC3-II. Downregulation of autophagy-specific protein Atg7 occurs in curcumin-treated HuT cells.

Taken together, our results indicated that curcumin by disrupting Hsp90 also disrupts important cellular pathway, autophagosome formation, in a ROS-dependent manner.

Cell proliferation and cell survival are regulated by a complex interactive network of cell signaling pathways. Therefore, it may not be sufficient to control malignant cell growth simply by disrupting 1 or 2 cellular targets. Research has established that one common biochemical change in malignant cells is the increased production of ROS due to high metabolism This fact is used to develop therapeutics against malignant cells.

Curcumin successfully uses this strategy to induce oxidative stress in HuT cells, perturb important cell survival mechanisms, and thus achieve high degree of killing.

Acquisition of data provided animals, acquired and managed patients, provided facilities, etc. Analysis and interpretation of data e. Administrative, technical, or material support i. This work was supported by Council of Scientific and Industrial Research CSIR , India, grants NWP and OLP First author M.

Khan and second author S. We suggest that while the fibroblast is a high value target for therapy, the use of small molecule intervention should consider the surrounding epithelium as a secondary adjunct target. N-acetylcysteine NAC , a precursor to the major antioxidant glutathione Demedts et al.

However, in clinical trials, adding NAC to the standard of care in IPF resulted in mixed findings and failed to produce significant evidence that NAC alone improves lung function in mild to moderately impaired patients Martinez et al.

Most disturbingly, in , a well-publicized clinical trial combining Prednisone, Azathioprine, and NAC in patients with IPF was prematurely stopped due to the adverse effects seen in the three-drug combination as compared to control Network, Yet, even in this trial, the conclusion drawn for NAC was mixed; the patients taking NAC alone were allowed to complete the trial fully as they did not demonstrate the adverse effects seen in the combination arm Martinez et al.

Curcumin is a hydrophobic polyphenol and the main active component of the spice turmeric. It has been used for thousands of years in Asian countries and traditional Ayurvedic medicine to inhibit scar tissue formation in open wounds Gupta et al. A significant application for curcumin in modern medicine has been highly elusive in spite of the large amount of effort and interest over the last half century Gupta et al.

While often characterized as having broad biological activities that can be applied to multiple diseases, the lack of significant translational success may be due to our poor understanding of the molecular mechanism.

Curcumin has also been reported to have anti-fibrotic capabilities in studies of wound healing, liver fibrosis and lung fibrosis models Lin et al. At the molecular level, curcumin has been reported to play an anti-fibrotic role by modulating transcription factors such as transforming growth factor beta Chen et al.

Due to these antifibrotic properties of curcumin, a number of investigators have hypothesized that this compound could serve as a possible therapeutic for IPF Smith et al. These studies focus on the antifibrotic effects of curcumin and, like many other in-vitro studies, report a reduction in profibrotic responses when pulmonary fibroblasts are treated in isolation.

However, these same studies report little significant improvement in bleomycin mouse models after curcumin treatment. We have previously observed that treatment of fibroblasts in vitro with curcumin induces both an increase in reactive oxygen species ROS production Rodriguez et al.

We hypothesize that increased oxidative stress may be a contributor to the pro-apoptotic properties of curcumin, and that alveolar epithelial cells may manage this burden more effectively possibly indicating that in the lung, curcumin is a fibroblast-specific drug.

We further suggest that if curcumin-induced apoptosis is through ROS production, then co-treatment with a potent ROS scavenger may inhibit apoptosis. To explore this hypothesis, we investigated the effects of curcumin and NAC on IPF derived pulmonary fibroblasts and epithelial cells in vitro.

IPF lung tissue was obtained through Inova Fairfax Hospital VA. All normal control lungs were obtained through the Washington Regional Transplant Community WRTC.

Appropriate written informed consent was obtained for each patient and donor by Inova Fairfax hospital and the WRTC. This study was approved by the Inova Fairfax Hospital Internal Review Board IRB All experiments were performed in accordance with relevant guidelines and regulations. The primary fibroblasts used in this study were isolated from human lungs procured in the operating room within minutes of explantation.

The lungs were oriented from apex to base, and all samples used in this study were taken from the peripheral lower lobe of the lung.

Fibroblasts were isolated from the lung tissue of four patients with advanced IPF IPF-F and four normal lungs NHLF using differential binding. Differential binding applied in this study is a modified protocol from that previously described Emblom-Callahan et al.

The attached fibroblast population was then vigorously washed with PBS to remove any unattached cells. The resuspended pellet containing primary epithelial cells was transferred to tissue culture plastic and placed into the incubator for continued culture. Analysis was carried out on cells within the 2—5 passage range.

QPCR was carried out using Quantifast SYBR Green PCR Kit Qiagen. QPCR was carried out in triplicate using mRNA specific primers Table 1 and normalized to 18S expression levels using the delta-delta CT method Pfaffl, To assay the in-vitro cell migration capability of each fibroblast cell line we used a modified protocol based on methodologies published by Liang et al.

Liang et al. Cells were then seeded in a Costar® 6-well tissue culture treated plate at a concentration of , cells per well. Images of the scratch were captured at 4X magnification using an EVOS FL Auto Light Microscope Life Technologies. The scratch test images were analyzed using TScratch Version 1.

Prior to each experiment, all cells were pre-treated in the same manner. Cells were then seeded in triplicate at cells per well in a 96 well plate in complete media and allowed to attach overnight. Cells were then challenged for h in varying concentrations of curcumin, NAC, or co-treatment as reported in the results section.

Quantification of cell number was performed using the CellTiter 96® AQ ueous Cell Proliferation Assay Promega. Statistical analysis was performed with Microsoft Excel using student t-tests.

A corresponding P -values of less than 0. To confirm the observation that curcumin can inhibit the myofibroblast phenotype in pulmonary fibroblasts Smith et al. We observe significant decreases in the expression of myofibroblast activation and proliferation associated genes in both the NHLF and IPF-F Fig.

Initial analysis of the data seemed to indicate that the antifibrotic effect of curcumin on IPF-F was more pronounced as the gene expression reduction was greater in IPF-F. However, this was not statistically significant and is likely attributable to the significant heterogeneity in the gene expression profile observed in IPF-F as compared to NHLF Fig.

We next explored the effect of NAC on the antifibrotic effect of curcumin Fig. The co-administration of curcumin and NAC continued to demonstrate a reduction in Smooth Muscle Actin ACTA2 and Proliferating Cell Nuclear Antigen PCNA in IPF-F as compared to untreated control, however the expression of Collagen 1A1 COL1A1 and Cyclin D CCND1 was not significantly altered.

This effect was not observed in the NHLF. This significant trend is also observed for PCNA in NHLF. To assess the functional effect of curcumin on the migratory capability of the fibroblast we performed a h scratch test on IPF-F and NHLF Figs.

We report a significant decrease in the wound closure rate of both IPF-F and NHLF in the presence of curcumin alone.

Interestingly we also note that NAC alone reduced the migratory capability of NHLF Fig. However, the co-treatment of normal and IPF-F with NAC and curcumin does not inhibit the migratory capacity of the fibroblasts to the same degree as curcumin alone.

In fact, we observe no significant change in wound closure for NHLF in the presence of the co-therapy as compared to control. There remains, however, an attenuated migratory inhibition on IPF-F in our co-therapy.

Morphological changes observed in the curcumin treatment are also attenuated in the combination treatment. We, and others, have previously observed dose dependent increase in fibroblast apoptosis in the presence of curcumin Zhang et al.

We test a sublethal, high dose curcumin treatment to evaluate effect on epithelial cell viability Fig. After h curcumin exposure, we observed a significant reduction in NHLF and IPF-F viability but did not see a significant reduction in either A or primary epithelial cell viability.

Co-treatment of all cell populations with curcumin and NAC had no significant effect on cell viability. Finally, we observed, no significant changes of p21 or p53 expression in any cell type in the NAC or co-treatment group.

To asses this we measured induced reactive oxygen species ROS in our cells after exposure to curcumin, NAC, and combination of the two Fig. Interestingly, combined treatment with NAC and curcumin resulted in levels of ROS that are significantly reduced in all populations as compared to uninduced controls.

In addition to the generation of ROS species, we also report the changes in an oxidative stress response gene panel consisting of Hypoxia Inducible Factor 1α HIF1 , Superoxide Dismutase 2 SOD2 , Catalase CAT , and Nuclear factor-like 2 NRF2.

After curcumin challenge we observed significantly decreased expression of nearly all genes of this panel in both IPF-F and IPF epithelial cells. In NHLF we also see a significant reduction in expression of HIF1 and SOD2, but no concurrent change in NRF2 and catalase.

A gene expression deviates from this pattern with significant increases in expression of HIF1, NRF2, and catalase. In contrast with these results, the challenge of these cells with NAC alone resulted in decreased expression of one gene, SOD2, in the epithelial cells only.

Finally, combination treatment resulted in significant decreases in this same gene, SOD2, across all cell types with no change to any other gene within our panel. Expression of Catalase CAT is decreased in IPF derived fibroblasts while Nuclear Factor erythroid 2-related factor 2 NRF2 expression is decreased in IPF-F and epithelial cells.

In this study we sought to further explore the antifibrotic potential of curcumin and the effects that this molecule has on the IPF epithelial cell. We initially focused on the effects of curcumin on the myofibroblast phenotype to confirm previous studies and to establish a baseline from which to compare our later experiments.

We observed that hr treatment with curcumin was effective at reducing both the expression of myofibroblast associated genes such as COL1A1 and PCNA Fig. This observation confirmed previous reports Lin et al.

These data, coupled with recent fibroblast studies reporting curcumin dose dependent increases in apoptotic markers Zhang et al. With oxidative stress as a major stressor in all cells in the IPF lung, we included both epithelial cells and fibroblast in our investigations.

Measurement of ROS generation in these cells after curcumin challenge demonstrated a significant induction of ROS within the primary cells, however the immortal alveolar type II cell line A proved resistant Fig.

In addition to this, we noted a decrease in cell viability in the primary cells that was absent in A Fig. These data indicate that increased oxidative stress induced by curcumin may be activating an apoptotic cascade in the primary cells. In support of this hypothesis, we note that gene expression of the DNA damage response protein p53 is increased in all cell lines.

Concurrently, the transcriptional target of p53, and an S phase regulator - p21, is also increased in all cells.

A cells do seem to contradict our hypothesis as the increase in this same gene expression is not accompanied by apoptosis. We suggest that the high basal expression of the p53 inhibiting protein MDM2 in A cells may be the source of this discrepancy Liu et al.

Given our interest in IPF, we did not further explore this A pathway and rather, chose to alleviate the oxidative stress through NAC co-treatment.

To our surprise, at 40uM curcumin dosage in combination with NAC did not induce apoptosis Fig. In addition to this finding we also observed that the co-treatment inhibited NAC induced apoptosis in epithelial cells Fig.

Our hypothesis that reduction in curcumin induced oxidative stress would prevent apoptosis was supported by both the viability study and the reported reduction in ROS after NAC co-treatment Fig.

With these data in mind, we were interested in the genetic regulation of oxidative stress response genes in the presence of our two small molecules. When challenged with curcumin alone, all primary cell lines demonstrated a decreased expression of our oxidative stress panel Fig.

As with many other experiments, A proved an outlier with an increase in three of the four selected genes. Given our findings that curcumin induces ROS generation in these cells, these data indicate that the increased oxidative stress burden in primary cells is not accompanied by the robust response necessary to manage said burden.

The addition of NAC co-treatment did not have a significant effect on the expression of most genes in the panel but given that this co-treatment did alleviate ROS generation we did not expect to see an increase in these genes.

The one exception to this was the expression of SOD2. Our data indicates that SOD2 gene expression is tightly regulated by ROS generation. Given that this is the oxide dismutase found primarily in the mitochondria and, that mitochondria are a primary site for ROS generation, these data were complimentary.

A driving rationale for applying curcumin in IPF was the hypothesis that, in the lung, curcumin may be a fibroblast specific compound. Our study determined this to be somewhat untrue, but we suggest that our findings begin to develop a new paradigm for the application of curcumin and NAC in IPF.

We report that, as is the case for many antioxidant molecules Garry et al. In primary epithelial cells and fibroblasts, curcumin inhibits the oxidative stress response, while in As curcumin induces a strong oxidative stress response. However, it is also clear that this stress response is, in part, the result of the capacity of As to deal with an increased ROS burden.

Finally, these data reinforce that NAC is effective in reducing oxidative stress in pulmonary cells which is of significant therapeutic value where ROS production is induced. We note that the concentration of both curcumin and NAC used in this study are at high in-vitro concentrations.

Given the relatively low bioavailability of these compounds this is a significant concern in future translational studies. However, the clinical history of both compounds demonstrates that NAC and curcumin are well-tolerated compounds that can be given at high dosages without major secondary complications Gupta et al.

We also acknowledge that a weakness to our study lies in our lack of data on varied concentrations in the co-treatment. Further studies will focus on varying concentrations of these compounds and investigating modifications or alternatives that may increase the bioavailability of this therapy such as the application of the NAC sister drug NACA Aldini et al.

These findings indicate that the application of curcumin alone is an ineffective treatment option for use in IPF. The apoptotic effect of induced ROS is of significant concern in IPF, particularly given the high levels of oxidative stress already found in patient lungs.

Conversely, the alleviation of oxidative stress through NAC therapy alone is not a sufficient therapeutic approach. Our findings indicate that it may be possible to use these two treatments in combination, to elicit both the antifibrotic response and protect the surrounding epithelium from ROS induced apoptosis Fig.

The co-treatment of our cells with NAC did mitigate the antifibrotic potential of curcumin and; it is possible that increased concentrations of curcumin may overcome this attenuation, however it is just as likely that this will result in overwhelming ROS generation.

We suggest that further investigation into the oxidative stress generated by curcumin in pulmonary cells may aid in the elucidation of key pathways that can be manipulated to inhibit apoptosis and maintain high antifibrotic potential. Hypothetical Molecular Model of Curcumin and NAC Co-Treatment in IPF: Curcumin induces ROS mediated apoptosis in myofibroblasts releases excess ROS into the microenvironment.

As the IPF lung is an oxidative stress rich environment the excess ROS further damages epithelial cells in the lung. This propagates the wound healing response and may further induce fibrosis in a classical IPF feedforward loop.

The introduction of NAC co-treatment attenuates fibroblast apoptosis and alleviates ROS induced oxidative stress in epithelial cells. In turn this prevents additional fibroblast recruitment. Deduction of optimal in-vivo co-treatment concentrations may result in significant antifibrotic potential for therapeutic application.

In conclusion, the heterogeneity of IPF presents a significant challenge in the discovery of novel therapeutic approaches. Our findings suggest a novel combination of two molecules that have alone demonstrated a capacity to alleviate elements of the disease processes found in IPF.

Perhaps the key to treating IPF is not to strongly inhibit a single disease process, but instead attempt to alleviate multiple aberrant pathways through drug combinations.

Aggarwal BB, Sung B. Pharmacological basis for the role of curcumin in chronic diseases: an age-old spice with modern targets. Trends Pharmacol Sci. Article CAS Google Scholar. Aldini G, Altomare A, Baron G, Vistoli G, Carini M, Borsani L, et al.

N-acetylcysteine as an antioxidant and disulphide breaking agent: the reasons why. Free Radic Res. Amara N, Goven D, Prost F, Muloway R, Crestani B, Boczkowski J. Article Google Scholar.

Bando M, Hosono T, Mato N, Nakaya T, Yamasawa H, Ohno S, et al. Long-term efficacy of inhaled N-acetylcysteine in patients with idiopathic pulmonary fibrosis.

Intern Med. Behr J, Maier K, Degenkolb B, Krombach F, Vogelmeier C. Antioxidative and clinical effects of high-doseN-acetylcysteine in Fibrosing Alveolitis. Am J Respir Crit Care Med. Bui S. The Investigations of the Effect of In-Vitro Combination Treatment: Curcumin, Aspirin, and Sulforaphane on Idiopathic Pulmonary Fibrosis.

George Mason University; Accessed 28 Dec Camelo A, Dunmore R, Sleeman MA, Clarke DL. The epithelium in idiopathic pulmonary fibrosis: breaking the barrier. Front Pharmacol. Chen A, Zheng S. Curcumin inhibits connective tissue growth factor gene expression in activated hepatic stellate cells in vitro by blocking NF-κB and ERK signalling.

Br J Pharmacol. Chen W-C, Lai Y-A, Lin Y-C, Ma J-W, Huang L-F, Yang N-S, et al. J Agric Food Chem. Das L, Vinayak M. Long-term effect of curcumin down-regulates expression of tumor necrosis factor-α and interleukin-6 via modulation of E26 transformation-specific protein and nuclear factor-κB transcription factors in livers of lymphoma bearing mice.

Leuk Lymphoma. Datta A, Scotton CJ, Chambers RC. Novel therapeutic approaches for pulmonary fibrosis. Demedts M, Behr J, Buhl R, Costabel U, Dekhuijzen R, Jansen HM, et al. High-dose acetylcysteine in idiopathic pulmonary fibrosis.

N Engl J Med. Emblom-Callahan MC, Chhina MK, Shlobin OA, Ahmad S, Reese ES, Iyer EPR, et al. Genomic phenotype of non-cultured pulmonary fibroblasts in idiopathic pulmonary fibrosis. Garry A, Edwards DH, Fallis IF, Jenkins RL, Griffith TM. Ascorbic acid and tetrahydrobiopterin potentiate the EDHF phenomenon by generating hydrogen peroxide.

Cardiovasc Res. Gebäck T, Schulz MMP, Koumoutsakos P, Detmar M. Short technical reports. Gupta SC, Patchva S, Aggarwal BB. Therapeutic roles of curcumin: lessons learned from clinical trials.

AAPS J. Gupta SC, Patchva S, Koh W, Aggarwal BB. Discovery of curcumin, a component of the Golden spice, and its miraculous biological activities. Clin Exp Pharmacol Physiol. Hewlings SJ, Kalman DS. Hua Y, Dolence J, Ramanan S, Ren J, Nair S. Bisdemethoxycurcumin inhibits PDGF-induced vascular smooth muscle cell motility and proliferation.

Mol Nutr Food Res. Hutchinson JP, McKeever TM, Fogarty AW, Navaratnam V, Hubbard RB. Increasing global mortality from idiopathic pulmonary fibrosis in the twenty-first century. Ann Am Thorac Soc. Jha NS, Mishra S, Jha SK, Surolia A.

Antioxidant activity and electrochemical elucidation of the enigmatic redox behavior of curcumin and its structurally modified analogues. Electrochim Acta. Liang C-C, Park AY, Guan J-L. In vitro scratch assay: a convenient and inexpensive method for analysis of cell migration in vitro.

Nat Protoc. Lin Y-L, Lin C-Y, Chi C-W, Huang Y-T. Study on antifibrotic effects of curcumin in rat hepatic stellate cells. Phytother Res. Liu D, Gong L, Zhu H, Pu S, Wu Y, Zhang W, et al.

Molecular Medicine andd 25Xnd number: 27 Low glycemic lifestyle tips this Oxidatkve. Metrics details. Curcjmin Pulmonary Fibrosis Proper fueling before a sports tournament is a fatal lung disease of unknown etiology with only two federally approved drug options. Given the complex molecular pathogenesis of IPF involving multiple cell types and multiple pathways, we explore the effects of a potential antifibrotic and antioxidant drug combination. Curcumin is a polyphenolic compound derived from turmeric with significant biological activity including a potential antifibrotic capacity. Mohammad Aslam KhanSatindra Gahlot Oxiative, Sekhar Majumdar; Curcumin and Oxidative Stress Stress Performance enhancing drinks by Curcumin Promotes the Death Curcuimn Cutaneous T-cell Lymphoma HuT by Disrupting the Function of Several Molecular Targets. Mol Proper fueling before a sports tournament Curckmin 1 Sttress ; 11 9 : — Curcumin is known to exert its anticancer effect either by scavenging or by generating reactive oxygen species ROS. In this study, we report that curcumin-mediated rapid generation of ROS induces apoptosis by modulating different cell survival and cell death pathways in HuT cells. Curcumin induces the activation of caspase-8, -2, and -9, alteration of mitochondrial membrane potential, release of cytochrome cand activation of caspase-3 and concomitant PARP cleavage, but the addition of caspase inhibitors only partially blocked the curcumin-mediated apoptosis.

Author: Goltilrajas

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