Disclaimer: While this post has a sponsor, all facts and opinions expressed are those of the research article I am presenting as well as my own, none of which have been influenced in any way, shape, or form by the sponsor. Â Enjoy!
Fungicides are used throughout agriculture, and are certainly no stranger to viticulture / winegrowing.Â Â They are utilized in the vineyard to help protect the grapes against fungal attack, which is more common in more humid environments than in drier environments.Â One major concern regarding the use of fungicides, which in part has contributed to the organic viticulture movement, is the unintended environmental and human health impacts of the use of synthetic fungicides in the vineyard.Â Studies have found significant health impacts related to the use of conazole fungicides, including thyroid tumors and potentially other cancers.Â It is thought that these fungicides are related to higher incidences of cancerous tumors as a result of their endocrine disruption properties.Â In fact, endocrine disrupting compounds have been shown in hundreds (if not thousands) of studies to cause significant health problems in both wildlife and humans, including but not limited to reproductive health issues, neurological problems, and cancer.
One particular type of conazole fungicide that is used almost exclusively in vineyards is calledâ€śpenconazoleâ€ť.Â In 2008, the Commission of European Communities established a maximum amount of penconazole allowed to be used in vineyards, with the Maximum Residue Level reaching no higher than 0.2ppm for both table and wine grape vineyards.Â Studies have shown that many endocrine disruptors can still cause significant health problems at these very low doses, and that these established maximums may not be appropriate for curtailing negative health impacts in humans or wildlife.
The purpose of todaysâ€™ study, which is currently in press in the journal Toxicology in Vitro, was to look at genetic changes in human cells as a result of penconazole use in grapes, specifically looking at those genes that undergo a â€śchange in behaviorâ€ť after exposure of the fungicide.Â This study also aimed to determine if any of these genes could be used as indicators of penconazole exposure, which could be helpful in determining a treatment plan early and well ahead of any major carcinogenic tumor growth, as the prognosis for many cancers is almost certainly much better if the tumors are caught at a very early stage.
The cells used in this study were the human T-47D cell line, which had been originally isolated from a woman with an infiltrating ductal carcinoma (i.e. tumor) of the breast.Â These cells were used for several reasons: 1) because they are a human cell line; 2) because these cells possess several of the receptors required for endocrine activity (and thus endocrine disruption when applicable), including the estrogen receptor, androgen receptor, and progesterone receptor; and 3) because this cell line has been already studied and shown to be sensitive to endocrine disruptor in laboratory tests.
The fungicide penconazole was isolated from 50g of grapes that had been treated twice with the compound during the growing season. (Note: the paper did not indicate any more details than this in terms of the conditions in the vineyard).
Sensitivity of the cells was determined using the MTT assay, which basically gives information as to the percent survival of the cells after exposure to a toxin (in this case, penconazole).Â Penconazole was isolated from the grapes in concentrations between 0.1uM and 200uM.
Total RNA was isolated and analyzed to determine what (if any) genes were affected by the penconazole treatment.
The program Pathway-Express was employed to analyze the genes found to be activated to determine what the final outcome of this activation would be if complete.Â In other words, if a certain gene were activated due to penconazole exposure, the Pathway-Express program takes this information and basically says â€śhey, I see this gene is activatedĂ looks like you could end up with prostate cancer based on which gene is activated!â€ť.
- Penconazole exposure did not inhibit cell growth at the 0.1 and 1uM concentrations.
- Penconazole exposure at 10uM resulted in a 15% reduction in cell growth.
- Penconazole exposure resulted in the activation/modulation (or in other words, regulation of gene expression) of 2274 genes, with 507 of those genes being modulated in a similar fashion (and thus used for further analysis).
- Using the Pathway-Express program to analyze these 507 activated/modulated genes after penconazole exposure, the only significant pathway found was that of thyroid cancer.
- In other words, based on the genes that were activated after penconazole exposure, the end result (if the cells were actually in a human body) would be thyroid cancer.
- This pathway included 6 different genes: 2 down-modulated and 4 up-modulated genes.
- Down-modulated genes: Cyclin D1 (CCND1) and nuclear receptor coactivator 4 (NCOA4, a.k.a. PTC3)
- Cyclin D1 is involved in the regulation of CDK kinases, in particular the regulation of CDK4 and CDK6, which are critical for cellular growth processes.
- NCOA4 interacts with the androgen receptor in such a way to increase transcriptional activity (i.e. increase the protein-making process).
- These two genes were down-regulated, so in other words, they werenâ€™t as active as they would be under normal conditions.
- Up-modulated genes: retinoid X receptor alpha (RXRA), transcription factor 7-like 1 (TCF7L1), tropomyosin 3 (TPM3, a.k.a. TRK), and the translocated promoter region (TPR).
- RXRA codes for steroid and thyroid hormone transcriptional regulators, or in other words, is responsible for producing steroid and thyroid hormones.
- TCF7L1 functions in the regulation of cell cycle genes and cell death.
- TPM3 and TPR encode for the proteins that provide stability to the structurally stabilizing filaments of the cells, as well as for other proteins related to actin-binding.
- These four genes were up-regulated, so in other words, they are overexpressed and are producing a little too much of what they are responsible for.
Unless youâ€™ve recently had a class in developmental biology or genetics of some sort, a lot of what I just presented to you might sound a little confusing.Â Completely understandable!Â This isnâ€™t an easy topic to grasp, but once you are able to associate the more confusing terms with a more simplified approach, it doesnâ€™t look as foreboding.
Letâ€™s take a step back and review the main finding of this research, and then break it down from there.
TheÂ primary result from this research was that a human cell line exposed to the fungicide penconazole, which was isolated from grapes that had been exposed to the compound during the growing season, had notably altered gene expression such that thyroid cancer would be the most significant outcome.Â Simply put: exposure to the endocrine-disrupting fungicide penconazole increases the chances of getting thyroid cancer.Â This result is in agreement with other studied examining the effects of fungicides and pesticides on human health.
â€śOK, pretend I donâ€™t have a clue about this sort of thing: what the hell do you mean by all of this?â€ť
From the detailed genetic information found in this research, how is it that thyroid cancer is a possibility after exposure of human cells to penconazole?
Well, 6 genes were found to be related to this pathway, and that they were all significantly altered in the cells.Â Two of the genes were down-regulated, while 4 of the genes were up-regulated.Â Down-regulated in very basic terms means that the cells arenâ€™t producing as much as they would be under normal conditions, and up-regulated in very basic terms means that the cells are producing too much of what they would be under normal conditions.
So, in this case, two genes were malfunctioning after fungicide exposure to result in a poor showing in the cells, while four genes were malfunctioning after fungicide exposure to result in them going crazy with producing way too much â€śstuffâ€ť.
In this situation, the two genes that are down-regulated are ones that help regular cell division and proliferation.Â After exposure to penconazole, these genes get very â€śconfusedâ€ť and they are no longer able to effectively tell the cell where it is in the cell division and proliferation process, which can result in the process spinning out of control.Â Additionally, when you throw in the 4 up-regulated genes after penconazole exposure, you get even more cellular division and proliferation mayhem, as the cell no longer has the ability to stop producing steroid and thyroid hormones, AND it basically tells the cell to keep growing MORE.Â This sort of information (plus a lot more details) is what the Pathway-Express program was using to determine the ultimate outcome from this sort of down- and up-regulation, and with confidence it determined that these genes, when â€śmessed upâ€ť by the exposure to the fungicide penconazole, could result in thyroid cancer in whoever was initially exposed to the toxic compound.
According to the authors of this research, any of the genes that were shown to be affected by penconazole exposure, even at relatively low doses, could be possible â€śbiomarkers of penconazole exposureâ€ť.Â This could be an important tool, as it would identify possible at-risk individuals for thyroid cancer prior to the formation of any tumor, and perhaps preventative measures could be taken to attempt to reduce this risk, and at the very least monitor more frequently so that if a tumor were to appear later on after penconazole exposure, it could be found very early and effectively treated.
It is important to note who might be at-risk for exposure to penconazole, as well as other possible avenuesthat werenâ€™t tested in this study but would be important to study in the future.Â The penconazole exposure in this study came from the grapes after they had been treated a couple of times with the fungicide during the growing season.Â I think the most obvious people at risk for this type of situation are the vineyard workers, particular those with direct and frequent contact with a lot of grapes.Â I doubt there is enough penconazole on one grape that an inquisitive visitor would sneak in order to elicit this pathway to thyroid cancer, but certainly those exposed to the compound on a more regular and long-term basis may be at the greatest risk.
What about drinking the wine made from these grapes?Â Could penconazole fungicide exposure to the grapes result in thyroid cancer in the person that drank the wine made from those grapes?Â Now, thatâ€™s a loaded question, and one I canâ€™t answer with certainty from this particular study.Â It is highly likely that during the winemaking process, the penconazole is cleaned off and diluted so much that levels are too low to show this type of negative health impact, BUT then again, a lot of endocrine disruptors have been shown to be effective even at teeny tiny doses, so I donâ€™t want to just out and say â€śitâ€™s fineâ€ť without seeing the evidence.Â Also, there is also the possibility of interactions between the penconazole and the other chemicals/compounds used in the winemaking process, which could result in either negative effects, or a negated effect.Â Perhaps the synergistic interaction between penconazole and a particular winemaking component makes the fungicide more harmful to human health.Â Or on the other hand, perhaps the synergy results in the â€śshutdownâ€ť of penconazole and it is no longer a threat.
Finally, thinking about the post we discussed not too long ago about resveratrol and it being a seemingly healthy compound that happens to have endocrine disrupting-like characteristics:Â could resveratrol compete with the penconazole and come out â€śon topâ€ť, resulting in the shutdown of the fungicide in the system?Â Does wine inherently protect itself again this sort of disruptor simply by the nature of its own natural antioxidative, anti-carcinogenic, and other important health-benefiting qualities?
Certainly, I think these results show cause for concern for those people actually handling the grapes themselves after they have been treated with the fungicide penconazole, at least concern enough to do a long-term clinical observational study of those workers exposed to the compound on a somewhat regular basis and how many of them end up with some form of thyroid or other cancer after a certain number of years.
As far as exposure from wine is concerned, I think the jury is still out, but certainly these results should prompt other researchers to take this next step, as we really wonâ€™t know the true answer until someone actually does the study.
Even though I covered a lot, I know I didnâ€™t cover all of it.Â Please feel free to leave your comments either on what Iâ€™ve presented here, or perhaps questions, comments, or concerns that popped up in your mind based on the results that you read.Â Letâ€™s have a healthy discussion about this!Â All are welcome!
Source: Perdichizzi, S., Mascolo, M.G., Silingardi, P., Morandi, E., Rotondo F., Guerrini, A., Prete, L, Vaccari, M., and Colacci, A. 2013. Cancer-related genes transcriptionally induced by the fungicide penconazole. Toxicology in Vitro, http://dx.doi.org/10.1016/j.tiv.2013.06.006.