Palmitic Acid And Stearic Acid Induce Apoptosis In Rat Testicular Leydig Cell

murdoc

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Saturated free fatty acids, palmitic acid and stearic acid, induce apoptosis by stimulation of ceramide generation in rat testicular Leydig cell - ScienceDirect

In men, obesity has generally been associated with reduced plasma testosterone levels and with elevation of the plasma free fatty acids (FFAs). In this study, we investigated the effects of saturated FFAs including palmitic acid (PA) and stearic acid (SA), and polyunsaturatedFFA arachidonic acid (AA) on the survival of rat testicular Leydig cell cultured in vitro. PA and SA markedly suppressed Leydig cell survival in a time- and dose-dependent manner. In contrast, AA stimulated the cell proliferation at 5–10 times of physiological concentration. The suppressive effect of PA and SA on cell survival was caused by apoptosis evidenced by DNA ladder formation and Annexin V-EGFP/propidium iodide staining of the cells. The apoptotic effect of PA was possibly mediated by ceramide generation because it could be completely blocked by ceramide synthase inhibitor fumonisin B1 and exogenous ceramide itself could directly induce apoptosis in vitro. Surprisingly, the apoptosis induced by PA could be partly prevented by AA. These results indicate that PA and SA induce apoptosis in testicular Leydig cells by ceramide production and these apoptotic effects may be a possible mechanism for reproductive abnormalities in obese men, and AA can partly prevent the apoptotic effect induced by saturated FFA.

Obesity is associated with reduced plasma testosterone and elevation of the plasma free fatty acids.
That is in line with Peat.

Than they used rat testicular Leydig cell cultured in vitro and exposed them to palmitic acid, stearic acid arachidonic acid. Not surprisingly AA caused cell proliferation. I think under certain circumstances abnormal cell proliferation is carcinogenic and this forum had made me believe that AA is carcinogenic and SA quite the opposite.

My biochemical understanding is limited, so I have no explanation for the apoptotic effect induced by saturated fatty acid. But given the fact that stearic acid is discussed in this forum because of its positive health effects in terms of visceral fat, CVD and cancer, it seems odd to blame PA and SA for the reproductive abnormalities in obese men. I think the conclusion from this study are pretty far fetched.

Can someone clarify on this?
 

Hans

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If you block apoptosis, the cells simply undergo necrosis, which is definitely not what you want.
 

X3CyO

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Maybe its just cause fats arent stored in the testicles? It sounds like its bad either way.

Maybe itd be better to change the cell membranes composition instead.

Itd be cool to have the full article
 

Jon

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If you block apoptosis, the cells simply undergo necrosis, which is definitely not what you want.

So I'm confused? Is this saying that fat cell death was occurring in the leydig cells? Why would they be painting that in a bad color then? Shouldn't that increase androgenic activity?

Edit: nvm I see they're saying leydig cells were dying. Why would this be happening though? Is it just because the cells were omitting the fatty acid from access to the mitochondria, and the unsaturated fat was basically forcing its way into a cell that would have otherwise omitted it?
 
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Hans

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So I'm confused? Is this saying that fat cell death was occurring in the leydig cells? Why would they be painting that in a bad color then? Shouldn't that increase androgenic activity?

Edit: nvm I see they're saying leydig cells were dying. Why would this be happening though? Is it just because the cells were omitting the fatty acid from access to the mitochondria, and the unsaturated fat was basically forcing its way into a cell that would have otherwise omitted it?
Cells are constantly renewed. They are broken down through apoptosis and then new ones are made. When this doesn't happen, cells become damaged and necrosis takes place. Then you end up with a lot of damaged cells and proliferation starts taking place.
In vitro studies are not normal obviously, so the cells can't regulate the amount of fats and glucose that the researchers dump on them.
So most likely under normal in vivo conditions, saturated fat will ensure normal cell renewal cycles, whereas PUFAs will inhibit this and cause proliferation.
 

Jon

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Cells are constantly renewed. They are broken down through apoptosis and then new ones are made. When this doesn't happen, cells become damaged and necrosis takes place. Then you end up with a lot of damaged cells and proliferation starts taking place.
In vitro studies are not normal obviously, so the cells can't regulate the amount of fats and glucose that the researchers dump on them.
So most likely under normal in vivo conditions, saturated fat will ensure normal cell renewal cycles, whereas PUFAs will inhibit this and cause proliferation.

That's kind of what I figured. Your explanation of the ETC keeps replaying in my head lol. I've learned not to trust in vitro studies. Thanks for taking the time to teach me! I've learned a lot from you.
 

Hans

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That's kind of what I figured. Your explanation of the ETC keeps replaying in my head lol. I've learned not to trust in vitro studies. Thanks for taking the time to teach me! I've learned a lot from you.
Hey no problem man. We're all in this learning and growing process together. I just enjoy helping and sharing where I can. :thumbsup:
 

rei

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I have read that coconut oil contains something that is anti-virility if used as lubricant and it gets to absorb on the scrotum, would love to see scientific sources to rule out it being lubricant industry propaganda.
 
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Hey no problem man. We're all in this learning and growing process together. I just enjoy helping and sharing where I can. :thumbsup:
+1 I also have learned from you. You have a knack of breaking the complexity down into simple language for those of us with some base knowledge and zero scientific background. Thank you again @Salmonamb
 

Travis

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@Travis care to chime in?

I think this is a hypoxic response to unphysiological conditions. The use of fusarium to block ceramide synthesis (Fig. 4) convincingly demonstrates apoptosis can be reduced down to ceramide cytotoxicity, with is capable of inducing apoptosis in double-digit micromolar concentrations. The average concentration of free palmitate in plasma is 758±80 μmol (Groop, 1989), making a person wonder 'how we could even have testicles at all?' The maximum concentration used in this study is only half that of free palmitate in the plasma (Fig. 1), yet this concentration had caused massive reductions in Leydig cell count:

fig1.png fig3.png fig6.png fig5.png

Assuming the apoptosis essentially reduces-down to ceramide cytotoxicity the nuclear factor-κB could be mechanistically-linked. Nuclear factor-κB is a transcription factor normally residing in the cytosol that is activated by H₂O₂, oxidizing it's reduced thiols into a disulfide bridge in the process. This collapses the transcription factor, ostensibly lowering its water-solubility thereby facilitating nuclear diffusion. Nuclear factor-κB is a five fire alarm for reactive oxygen species and hypoxia, transcribing iNOS to scavenge superoxide via peroxynitrite formation (Ȯ₂⁻ + ṄO = ONOO⁻). The fact that NF-κB has been shown to become activated by high levels of ceramide suggests that H₂O₂ is formed in response (Kitajima, 1996).

The fact that ceramide cytotoxicity is linked to NF-κB activation implies that p53 had been activated, which is also a H₂O₂-sensitive transcription factor. If NF-κB is an alarm bell—the 'first responder' stationed near the membrane—the nuclear p53 could be no other than a doomsday switch. Activation of p53 means H₂O₂ had reached the nucleus, collapsing the transcription factor via disulfide bridge in a similar manner as NF-κB and HIF-1. Activated p53 actively induces apoptosis by transcribing for caspases, proteolytic enzymes that disassemble the cell's protein skeleton. This is accompanied by an increase in Bax and a decrease of Bcl2, a change noted by Lu et al. after feeding Leydig cells exclusively medium-chained saturated fatty acids and pyruvate (Fig. 5). The pyruvate would likely catalyze the elongation of palmitate into oleate, the two fatty acids that compose ceramide. The minimum essential medium (MEM) used also had free L-serine, the one amino acid that composes the 'backbone' of ceramide. The fetal calf serum of course has phosphate, also used by Lu et al., this being the fourth constituent of ceramide.

'Ceramide-induced apoptosis is accompanied by accumulation of p53 followed by an increase of Bax and decrease of Bcl-2 levels. Inhibition of p53 expression with p53 antisense oligonucleotides inhibits apoptosis and prevents the increase in Bax and decrease in Bcl-2. Furthermore, pretreatment with p53 antisense oligonucleotides markedly inhibits the induction of caspase activity. These results suggest that p53 regulates the ratio Bcl-2/Bax and the expression/ activation of caspases during ceramide-induced apoptosis in SKN-SH cells.' Kim

I would assume that ceramide cytotoxicity is nonspecific, or lacks a receptor, and works merely to induce a hypersaturation of the cell membrane. I would assume that any molecule analogous to ceramide would have similar effects—be it a phosphatide, sulfatide, or glycolipid. Incorporation of ceramide in the cell membrane could reduce glucose flux, oxygen flux, and catalyze the formation of H₂O₂ via superoxide dismutase (Ȯ₂⁻ + Cu⁺ + 2H⁺ = H₂O₂ + Cu²⁺). Selenoamino acids attenuate peroxide-induced p53 activation by forming selenoenzyme glutathione peroxidase, the other enzyme besides catalase responsible for (electronically) reducing cytosolic H₂O₂ into water. These Leydig cells were likely maintained in low-selenium conditions: Minimum essential medium (MEM) lacks selenium completely, and that found in fetal calf serum (FCS) depends on the dietary selenium intake of cows.

Considering the sub-physiological concentrations of pyruvate and palmitate able to induce Leydig cell death, 'why then do we even have testicles at all?' The Tokelauans have efficiently reproduced under high-coconut conditions for generations, and judging by their phenotype they appear to have adequate androgen concentrations. The answer could lie with the sum total of free unsaturated fatty acids found in the plasma—i.e. oleate, α-linoleate, palmitoleate—acting to maintain the degree of cell membrane unsaturation required for glucose and oxygen flux; this is perhaps why arachidonate was able to 'increase survival' under these conditions (Fig. 6). If arachidonate is vitally acting in this manner, a person could confidently predict that both EPA or DHA would appear more protective under conditions used.

By feeding the Leydig cells sodium pyruvate, palmitate, phosphate, and L-serine: you'd suspect these researchers had intentionally concocted a recipe for ceramide synthesis. This relatively saturated ceramide phospholipid is normally produced with intent of myelinating nerves, along with with the steroids: cholesterol, pregnenolone, and progesterone. Ceramide is certainly not a normal membrane lipid for Leydig cells, but rather a myelin lipid that can only accumulate to apoptotic levels under unphysiological culture conditions or nerve damage.
 
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Jon

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I think this is a hypoxic response to unphysiological conditions. The use of fusarium to block ceramide synthesis (Fig. 4) convincingly demonstrates apoptosis can be reduced down to ceramide cytotoxicity, with is capable of inducing apoptosis in double-digit micromolar concentrations. The average concentration of free palmitate in plasma is 758±80 μmol (Groop, 1989), making a person wonder 'how we could even have testicles at all?' The maximum concentration used in this study is only half that of free palmitate in the plasma (Fig. 1), yet this concentration had caused massive reductions in Leydig cell count:

View attachment 10510 View attachment 10511 View attachment 10512 View attachment 10513

Assuming the apoptosis essentially reduces-down to ceramide cytotoxicity the nuclear factor-κB could be mechanistically-linked. Nuclear factor-κB is a transcription factor normally residing in the cytosol that is activated by H₂O₂, oxidizing it's reduced thiols into a disulfide bridge in the process. This collapses the transcription factor, ostensibly lowering its water-solubility thereby facilitating nuclear diffusion. Nuclear factor-κB is a five fire alarm for reactive oxygen species and hypoxia, transcribing iNOS to scavenge superoxide via peroxynitrite formation (Ȯ₂⁻ + ṄO = ONOO⁻). The fact that NF-κB has been shown to become activated by high levels of ceramide suggests that H₂O₂ is formed in response (Kitajima, 1996).

The fact that ceramide cytotoxicity is linked to NF-κB activation implies that p53 had been activated, which is also a H₂O₂-sensitive transcription factor. If NF-κB is an alarm bell—the 'first responder' stationed near the membrane—the nuclear p53 could be no other than a doomsday switch. Activation of p53 means H₂O₂ had reached the nucleus, collapsing the transcription factor via disulfide bridge in a similar manner as NF-κB and HIF-1. Activated p53 actively induces apoptosis by transcribing for caspases, proteolytic enzymes that disassemble the cell's protein skeleton. This is accompanied by an increase in Bax and a decrease of Bcl2, a change noted by Lu et al. after feeding Leydig cells exclusively medium-chained saturated fatty acids and pyruvate (Fig. 5). The pyruvate would likely catalyze the elongation of palmitate into oleate, the two fatty acids that compose ceramide.

'Ceramide-induced apoptosis is accompanied by accumulation of p53 followed by an increase of Bax and decrease of Bcl-2 levels. Inhibition of p53 expression with p53 antisense oligonucleotides inhibits apoptosis and prevents the increase in Bax and decrease in Bcl-2. Furthermore, pretreatment with p53 antisense oligonucleotides markedly inhibits the induction of caspase activity. These results suggest that p53 regulates the ratio Bcl-2/Bax and the expression/ activation of caspases during ceramide-induced apoptosis in SKN-SH cells.' Kim

I would assume that ceramide cytotoxicity is nonspecific, or lacks a receptor, and works merely to induce a hypersaturation of the cell membrane. I would assume that any molecule analogous to ceramide would have similar effects—be it a phosphatide, sulfatide, or glycolipid. Incorporation of ceramide in the cell membrane could reduce glucose flux, oxygen flux, and catalyze the formation of H₂O₂ via superoxide dismutase (Ȯ₂⁻ + Cu⁺ + 2H⁺ = H₂O₂ + Cu²⁺). Selenoamino acids attenuate peroxide-induced p53 activation by forming selenoenzyme glutathione peroxidase, the other enzyme besides catalase responsible for (electronically) reducing cytosolic H₂O₂ into water. These Leydig cells were likely maintained in low-selenium conditions: Minimum essential medium (MEM) lacks selenium completely, and that found in fetal calf serum (FCS) depends on the dietary selenium intake of cows.

Considering the sub-physiological concentrations of pyruvate and palmitate able to induce Leydig cell death, 'why then do we even have testicles at all?' The Tokelauans have efficiently reproduced under high-coconut conditions for generations, and judging by their phenotype they appear to have adequate androgen concentrations. The answer could lie with the sum total of free unsaturated fatty acids found in the plasma—i.e. oleate, α-linoleate, palmitoleate—acting to maintain the degree of cell membrane unsaturation required for glucose and oxygen flux; this is perhaps why arachidonate was able to 'increase survival' under these conditions (Fig. 6). If arachidonate is vitally acting in this manner, a person could confidently predict that both EPA or DHA would appear more protective under conditions used.

By feeding the Leydig cells sodium pyruvate and free palmitate, yet no other, these researchers had given them a veritable recipe for ceramide synthesis. This relatively saturated phospholipid is normally produced with intent of myelinating nerves. along with: cholesterol, pregnenolone, and progesterone. This is certainly not a normal membrane lipid for Leydig cells, and which can only accumulate to apoptotic levels under unphysiological culture conditions.

Just can't trust those in vitro studies. Thank you for the thourough opinion/explanation :) so much of the learnings I have received as @Amazoniac might say!
 

Wagner83

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[...]which can only accumulate to apoptotic levels under unphysiological culture conditions.
[Thanks]

Do you think applying saturated fats to the scrotum could replicate some of those in vitro effects? Ray had mentioned he wouldn't mess with the lipid composition of the testicles/scrotum and it sounds like studies like this could be one clue as to why.
 

dannibo

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Cells are constantly renewed. They are broken down through apoptosis and then new ones are made. When this doesn't happen, cells become damaged and necrosis takes place. Then you end up with a lot of damaged cells and proliferation starts taking place.
In vitro studies are not normal obviously, so the cells can't regulate the amount of fats and glucose that the researchers dump on them.
So most likely under normal in vivo conditions, saturated fat will ensure normal cell renewal cycles, whereas PUFAs will inhibit this and cause proliferation.
Pufas do not inhìbit any area of life. They are in fact life essence. Omega6 and Omega3 in a ratio of 2:1 approx are absolutely essential to functional life. The problem is "parroting" bull**** from guesses. Scientific papers by exceptional scientists have proven that cold pressed organic EFAs can and do reverse diseases including cancers and more importantly atherosclerosis.
 

Hans

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Pufas do not inhìbit any area of life. They are in fact life essence. Omega6 and Omega3 in a ratio of 2:1 approx are absolutely essential to functional life. The problem is "parroting" bull**** from guesses. Scientific papers by exceptional scientists have proven that cold pressed organic EFAs can and do reverse diseases including cancers and more importantly atherosclerosis.
With cold-pressed EFAs you're referring to specific oils or actually isolated linoleic and linolenic acid?
 

PaRa

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With cold-pressed EFAs you're referring to specific oils or actually isolated linoleic and linolenic acid?
probably just to not oxidized pufas

eggs, nuts, small fish, avocado seems to just be net positive in humans

and unsaturated fats lower LPS induced endotoxinemia contrary to SFA
and lowered endotoxinemia = lowered serotonin
 

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