How has our perspective of VC pathogenesis changed in recent years?
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Marwa Alm
– Medial calcification ( observed in CKD pts), was originally thought to be the result of passive depositions blood Ca and inorganic Pi.
– Two decades ago, genes and proteins normally restricted to bone tissue were found to be expressed in calcified arteries CKD pts.
– in CKD pts, Pi concentration increases while calcification inhibitors decreses e.g fetuin A, when this occurs: Pi & Ca aggregates forming amorphous soluble CPP1, which matures to crystalline CPP2, upregulating osteogenic proteins, preceeding osteogenic trans-differentiation with resultant VSMC.
-Recent insights regarding the impact of Mg on CPP maturation, expanded the spectrum of mechanisms that underlie the anti- calcifing properties of Mg.
– the intruduction of calcification propensity test T50 (which measures the invitro transition time from CPP1 to CPP2) has created an opportunity to quantify the mineral buffering capacity of CKD pts to withstand vascular calcification – Over the past decade, in pts on hemodialysis, a higher S.Mg has been associated with lower CV mortality risk. However too highly elevated blood Mg may compromise bone mineralization ( as Mg is known to interfere with hydroxyapatite formation
High phosphate and absence of calcification inhibitors e.g., Fetuin A: lead to formation of the amorphous primary calciprotein patricle (CPP1) in CKD patient (Aggregates of serum proteins, Ca/ Pi crystals).
CPP1 mature into CCP2 because of absence of crystallization inhibitors.
CCP2 & Pi induce calcification in VSMCs and increase expression of pro calcification genes e.g., RUNX2, BMP2 , OPN,ALP, & OSX(SP7). At the same time there is a decrease in contractile genes such as transgelin, The resulting calcified VSMCs amplify the calcification events by: shedding Ca – loaded exosomes which are then taken up ( engulfed) by the neighboring VSMCs
The experimental studies and recent information showed that the most important is the transition from CPP1 to CPP’. CPP formation is mainly dependent on calcium, Pi and fetuin-A concentration. The introduction of the calcification propensity test (T50 test) and the identification of magnesium as a targetable factor to modify CPP2 maturation provided us with an interesting tool regarding vascular calcification in CKD
As CPP formation mainly depends on Ca2þ, Pi and fetuin-A concentrations, CPP may explain Pi-stimulated vascular calcification in CKD and thus mediate Pi-induced VSMC calcification. Consequently these pathological effects of Pi may be reduced by preventing CPP2 formation by Mg2þ. Indeed, many experimental vascular calcification inhibitors were shown to delay CPP2maturation. The introduction of the calcification propensity test (T50 test), which measures the transition time from CPP1 to CPP2 ex vivo in serum samples, has created an opportunity to quantify the mineral-buffering capacity ofCKD patients to withstand vascular calcification . A higher calcification propensity (T50) is associated with mortality and the presence and severity of vascular calcification in patients with CKD. Factors reducing calcification propensity are ofinterest as potential treatment options for progressive vascular calcification. The identification of Mg2þ as a targetable factor to modify
CPP2 maturation provides an interesting clinical tool for vascular calcification in CKD. Increasing dialysate Mg2þ concentration from 0.5 to 1.0mmol/L led to an increase in serum Mg2þ concentration of 0.34mmol/L and improved T50 significantly . In another study, ex vivo addition of Mg2þ to serum samples of pre-transplant CKD patients, dosedependently improved T50 .In 2019, a clinical trial was published in which MgO was supplemented in 96 pre-dialysis CKD patients, leading to decreased progression of coronary artery calcification, although no effect was seen on thoracic aorta calcification. Now the stage is set for clinical trials in patients with CKD and on dialysis to assess the effectiveness and safety of Mg2þ supplementation in CKD patients on clinically relevant endpoints. However, proper monitoring of efficacy is challenging because establishing differences in clinically relevant endpoints requires long-term follow-up ofa substantial number of study participants. It is tempting to assume that, for instance, changes in the T50 antedate changes in these clinical endpoints. While T50 is an informative indicator ofcalcification propensity in CKD, it is of importance to determine whether Mg2þ supplementation ultimately results in lower calcification scores and its associated clinical events in CKD patients.
How has our perspective of VC pathogenesis changed in recent years?High Pi concentrations and the absence of circulating inhibitors such as fetuin-A stimulate the formation of CPP1 in the circulation of CKD patients. Subsequently CPP1 transitions into CPP2 due to a lack of circulating crystallization inhibitors. 2 – CPP2 and Pi induce calcification in VSMCs and stimulate expression of pro-calcification genes such as RUNX2, ALP (ALPL) and osterix (SP7).
Simultaneously, contractility genes such as transgelin (SM22a) diminish. 3 – Combined, this cascade results in VSMC transdifferentiation and calcification and loss of VSMC function and contractility. 4 – The resulting calcified VSMCs amplify the calcification process by shedding Caloaded exosomes that are engulfed by neighbouring VSMCs
How has our perspective of VC pathogenesis changed in recent years?
High Pi and absence of calcification inhibitors e.g., Fetuin A: lead to formation of the amorphous primary calciprotein patricle (CPP1) in CKD patient (Aggregates of serum proteins, Ca/ Pi crystals).
CPP1 mature into CCP2 because of absence of crystallization inhibitors.
CCP2 & Pi induce calcification in VSMCs and increase expression of pro calcification genes e.g., RUNX2, BMP2 , OPN,ALP, & OSX(SP7). At the same time there is a decrease in contractile genes such as transgelin, The resulting calcified VSMCs amplify the calcification events by: shedding Ca – loaded exosomes which are then taken up ( engulfed) by the neighboring VSMCs
vascular calcification is an active cell-orchestrated process by transdifferentiation of vascular smooth muscle cells (VSMCs) into osteoblast-like cells.
VMSCs are triggered by the uremic milieu, well established for Pi and indoxyl sulphate, to change their transcriptional repertoire and lose their contractile phenotype .
These transformed VSMCs are characterized by the expression of genes such as Runt-related transcription factor 2 (RUNX2), bone morphogenetic proteins (BMP), osteopontin (OPN encoded by SPP1) and osterix (OSX encoded by SP7), as well as higher alkaline phosphatase (ALP) activity ,thereby resembling osteoblasts.
These cells orchestrate the local spread of vascular calcification by increasing extracellular matrix synthesis and releasing Ca-rich exosomes .Experimental evidence strongly suggests that these processes are directly initiated at least partially by excess extracellular Pi and its increased cellular uptake .
Interestingly, the results of other studies suggest that VSMC transdifferentiation is a consequence rather than a cause of the vascular calcification process.
elevated blood Pi concentrations give rise to colloidal particles in the circulation.
In healthy individuals, despite supersaturated concentrations of soluble Ca and Pi in the circulation, the step from soluble Pi towards amorphous Ca–Pi rarely occurs, due to the presence of inhibitors (e.g. fetuin-A, albumin, osteopontin).
In CKD, the Pi concentration increases even more and the expression of inhibitory proteins or colloidal components of these particles, such as fetuin A, decreases .
Therefore in the circulation of CKD patients, this protective mineral-buffering system is compromised.
When this occurs, Pi, Ca and serum proteins aggregate to form amorphous, soluble Ca–Pi particles or primary calciprotein particles (CPP1).
Depending on the local milieu, CPP1 mature to crystalline secondary calcipro- tein particles (CPP2). The maturation time of CPP1–CPP2, which is measured ex vivo in serum samples in the recently developed T50 test, has been proposed as a measure for the calcification propensity in individuals .
CPP2 have been identified in the circulation of CKD patients and their presence is associated with aortic stiffness .
CPP2, in contrast to CPP1, have been shown to directly induce VSMC calcification in cultured VSMCs.
Accordingly, vascular calcification can conceptually be prevented by inhibiting hydroxyapatite formation despite persistently high extracellular Pi concentrations in VSMCs.
Altogether, these findings suggest that not Pi as such, but rather the formation of CPP2, is the true culprit in vascular calcification initiation and progression .
The pathogenesis of vascular calcification changes in recent years.
High Po4and absence of calcification inhibitors e.g., Fetuin A lead to formation of the amorphous primary calciprotein patricle in CKD patient
The resulting calcified VSMCs amplify the calcification events by shedding Ca – loaded exosomes which are then taken up ( engulfed) by the neighboring VSMCs
Also the recent study demonstrate the normal mg level decrease possiblity of vascular calcification via inhibiton of transdifferntiation of smooth muscle in blood vessels
1-High Pi concentrations and the absence of circulating inhibitors such as fetuin-A stimulate the formation of CPP1 in the circulation of CKD patients. Subsequently CPP1 transitions into CPP2 due to a lack of circulating crystallization inhibitors. 2 – In vitro studies suggest that CPP2 and Pi induce calcification in VSMCs and stimulate expression of pro-calcification genes such as RUNX2, ALP (ALPL) and osterix (SP7). Simultaneously, contractility genes such as transgelin (SM22a) diminish.
3 – Combined, this cascade results in VSMC transdifferentiation and calcification and loss of VSMC function and contractility.
4 – The resulting calcified VSMCs amplify the calcification process by shedding Ca- loaded exosomes that are engulfed by neighbouring VSMCs.
the pathogenesis of vascular calcification has 2 main mechanisms:
1- the transdifferentiation of vascular smooth muscle into osteoblast like cells
2- formation of secondary calciprotein particles
The medial calcification in Ckd occur due to the VSMC transdifferentiate to osteoblast like cell with expression of gene like RUNX, BMP, osteopontin, osx and alkaline phosphotase, these will cause increase of extracellular matrix and release of Ca from exosome, these initiated by increase phosphorus.
As phosphorous increase with Calcium and decrease inhibitor (fetuin A, albumin, osteopontin) so they combined to form amorphous soluble Ca-ph particle or primary CCP which mature to form secondary CCP which is form needle shape calcification in the medial layer of vessels which can be prevented by high Mg because once it occur cannot inhibit the process. The maturation time from CCP1 to CCP2 called T50 which is the measure of of calcification propensity in indiv
Recent studies have shown that magnesium can inhibit the maturation of calciprotein particles (CPPs), which are implicated in the development of vascular calcification.
Additionally, magnesium can protect the mineral-buffering system that is overwhelmed by phosphate in CKD, thereby reducing the risk of mineral deposition in the vasculature.
Magnesium also appears to play a role in preventing vascular smooth muscle cells (VSMCs) from transdifferentiating into an osteoblast-like phenotype, thereby preventing the expression of osteogenic genes and the production of calcification-promoting substances.
A clinical perspective of modifying Mg balance Recently as CPP formation depends on Ca, Pi, and futin-A concentration, CPP may explain Pi-stimulated vascular calcification in CKD and thus mediate Pi-induce VSMC calcification, so; Magnesium can prevent CPP2 formation by reducing Pi effect, and this supports the need to quantify the mineral-buffering capacity in CKD patients to withstand vascular calcification, and this may open a promising window toward a potential treatment target for vascular calcification. Meanwhile, measures have to be taken to reduce this risk;
In view of recent studies showing that CPP2 induces VSMC transdifferentiation and vascular calcification, the hypothesis indicating a prominent role for Mg2+ on intracellular pro-calcifying pathways may be insufficient.
CPP2 may develop before VSMCs become osteoblast-like. Mg2+ inhibition of CPP2 maturation will diminish osteogenic gene expression and keep VSMCs contractile.
Recent studies demonstrate that magnesium suppresses calciprotein particle formation, which may explain its anti-calcification effects. Magnesium protects the calcification milieu and restores the mineral-buffering system that phosphate overwhelms in CKD patients.
Magnesium is a regulator of calciprotein particle maturation and extracellular matrix mineralization, making it a viable new therapeutic strategy for treating CKD vascular calcification.
the evolution and the recent explanation and understanding the pathogenesis of vascular calcification in CKD and in dialysis pt.
and more study and research about new non traditional risk factors as role of Mg. Calcipotriene particle maturation , Pi, FGF 23 , …..other
will change our perspective in the pathogenesis of VC and promising novel treatment and practical guide lines to decrease risk VC and mortality
How has our perspective of VC pathogenesis changed in recent years? Three steps:
Initiation
Acceleration
Amplification
How does it happened ?
High Pi and absence of calcification inhibitors e.g., Fetuin A lead to formation of the amorphous primary calciprotein patricle (CPP1) in CKD patient (Aggregates of serum proteins, Ca/ Pi crystals).
Primary calciprotein particle (CPP1) mature into secondary calciprotein particle (CCP2) because of absence of crystallization inhibitors.
CCP2 & Pi induce calcification in VSMCs and increase expression of pro calcification genes e.g., RUNX2, BMP2 , OPN, ALP, & OSX(SP7). At the same time there is a decrease in contractile genes such as transgelin.
The resulting calcified VSMCs amplify the calcification events by shedding Ca – loaded exosomes which are then taken up ( engulfed) by the neighboring VSMCs
– Medial calcification ( observed in CKD pts), was originally thought to be the result of passive depositions blood Ca and inorganic Pi.
– Two decades ago, genes and proteins normally restricted to bone tissue were found to be expressed in calcified arteries CKD pts.
– in CKD pts, Pi concentration increases while calcification inhibitors decreses e.g fetuin A, when this occurs: Pi & Ca aggregates forming amorphous soluble CPP1, which matures to crystalline CPP2, upregulating osteogenic proteins, preceeding osteogenic trans-differentiation with resultant VSMC.
-Recent insights regarding the impact of Mg on CPP maturation, expanded the spectrum of mechanisms that underlie the anti- calcifing properties of Mg.
– the intruduction of calcification propensity test T50 (which measures the invitro transition time from CPP1 to CPP2) has created an opportunity to quantify the mineral buffering capacity of CKD pts to withstand vascular calcification
– Over the past decade, in pts on hemodialysis, a higher S.Mg has been associated with lower CV mortality risk. However too highly elevated blood Mg may compromise bone mineralization ( as Mg is known to interfere with hydroxyapatite formation
The experimental studies and recent information showed that the most important is the transition from CPP1 to CPP’. CPP formation is mainly dependent on calcium, Pi and fetuin-A concentration. The introduction of the calcification propensity test (T50 test) and the identification of magnesium as a targetable factor to modify CPP2 maturation provided us with an interesting tool regarding vascular calcification in CKD
As CPP formation mainly depends on Ca2þ, Pi and fetuin-A concentrations, CPP may explain Pi-stimulated vascular calcification in CKD and thus mediate Pi-induced VSMC calcification. Consequently these pathological effects of Pi may be reduced by preventing CPP2 formation by Mg2þ. Indeed, many experimental vascular calcification inhibitors were shown to delay CPP2maturation. The introduction of the calcification propensity test (T50 test), which measures the transition time from CPP1 to CPP2 ex vivo in serum samples, has created an opportunity to quantify the mineral-buffering capacity ofCKD patients to withstand vascular calcification . A higher calcification propensity (T50) is associated with mortality and the presence and severity of vascular calcification in patients with CKD. Factors reducing calcification propensity are ofinterest as potential treatment options for progressive vascular calcification. The identification of Mg2þ as a targetable factor to modify
CPP2 maturation provides an interesting clinical tool for vascular calcification in CKD. Increasing dialysate Mg2þ concentration from 0.5 to 1.0mmol/L led to an increase in serum Mg2þ concentration of 0.34mmol/L and improved T50 significantly . In another study, ex vivo addition of Mg2þ to serum samples of pre-transplant CKD patients, dosedependently improved T50 .In 2019, a clinical trial was published in which MgO was supplemented in 96 pre-dialysis CKD patients, leading to decreased progression of coronary artery calcification, although no effect was seen on thoracic aorta calcification. Now the stage is set for clinical trials in patients with CKD and on dialysis to assess the effectiveness and safety of Mg2þ supplementation in CKD patients on clinically relevant endpoints. However, proper monitoring of efficacy is challenging because establishing differences in clinically relevant endpoints requires long-term follow-up ofa substantial number of study participants. It is tempting to assume that, for instance, changes in the T50 antedate changes in these clinical endpoints. While T50 is an informative indicator ofcalcification propensity in CKD, it is of importance to determine whether Mg2þ supplementation ultimately results in lower calcification scores and its associated clinical events in CKD patients.
How has our perspective of VC pathogenesis changed in recent years?High Pi concentrations and the absence of circulating inhibitors such as fetuin-A stimulate the formation of CPP1 in the circulation of CKD patients. Subsequently CPP1 transitions into CPP2 due to a lack of circulating crystallization inhibitors. 2 – CPP2 and Pi induce calcification in VSMCs and stimulate expression of pro-calcification genes such as RUNX2, ALP (ALPL) and osterix (SP7).
Simultaneously, contractility genes such as transgelin (SM22a) diminish. 3 – Combined, this cascade results in VSMC transdifferentiation and calcification and loss of VSMC function and contractility. 4 – The resulting calcified VSMCs amplify the calcification process by shedding Caloaded exosomes that are engulfed by neighbouring VSMCs
How has our perspective of VC pathogenesis changed in recent years?
vascular calcification is an active cell-orchestrated process by transdifferentiation of vascular smooth muscle cells (VSMCs) into osteoblast-like cells.
VMSCs are triggered by the uremic milieu, well established for Pi and indoxyl sulphate, to change their transcriptional repertoire and lose their contractile phenotype .
These transformed VSMCs are characterized by the expression of genes such as Runt-related transcription factor 2 (RUNX2), bone morphogenetic proteins (BMP), osteopontin (OPN encoded by SPP1) and osterix (OSX encoded by SP7), as well as higher alkaline phosphatase (ALP) activity ,thereby resembling osteoblasts.
These cells orchestrate the local spread of vascular calcification by increasing extracellular matrix synthesis and releasing Ca-rich exosomes .Experimental evidence strongly suggests that these processes are directly initiated at least partially by excess extracellular Pi and its increased cellular uptake .
Interestingly, the results of other studies suggest that VSMC transdifferentiation is a consequence rather than a cause of the vascular calcification process.
Calciprotein particle formation mediate phosphate- induced calcification
elevated blood Pi concentrations give rise to colloidal particles in the circulation.
In healthy individuals, despite supersaturated concentrations of soluble Ca and Pi in the circulation, the step from soluble Pi towards amorphous Ca–Pi rarely occurs, due to the presence of inhibitors (e.g. fetuin-A, albumin, osteopontin).
In CKD, the Pi concentration increases even more and the expression of inhibitory proteins or colloidal components of these particles, such as fetuin A, decreases .
Therefore in the circulation of CKD patients, this protective mineral-buffering system is compromised.
When this occurs, Pi, Ca and serum proteins aggregate to form amorphous, soluble Ca–Pi particles or primary calciprotein particles (CPP1).
Depending on the local milieu, CPP1 mature to crystalline secondary calcipro- tein particles (CPP2). The maturation time of CPP1–CPP2, which is measured ex vivo in serum samples in the recently developed T50 test, has been proposed as a measure for the calcification propensity in individuals .
CPP2 have been identified in the circulation of CKD patients and their presence is associated with aortic stiffness .
CPP2, in contrast to CPP1, have been shown to directly induce VSMC calcification in cultured VSMCs.
Accordingly, vascular calcification can conceptually be prevented by inhibiting hydroxyapatite formation despite persistently high extracellular Pi concentrations in VSMCs.
Altogether, these findings suggest that not Pi as such, but rather the formation of CPP2, is the true culprit in vascular calcification initiation and progression .
How has our perspective of VC pathogenesis changed in recent years?
The pathogenesis of vascular calcification changes in recent years.
High Po4and absence of calcification inhibitors e.g., Fetuin A lead to formation of the amorphous primary calciprotein patricle in CKD patient
The resulting calcified VSMCs amplify the calcification events by shedding Ca – loaded exosomes which are then taken up ( engulfed) by the neighboring VSMCs
Also the recent study demonstrate the normal mg level decrease possiblity of vascular calcification via inhibiton of transdifferntiation of smooth muscle in blood vessels
1-High Pi concentrations and the absence of circulating inhibitors such as fetuin-A stimulate the formation of CPP1 in the circulation of CKD patients.
Subsequently CPP1 transitions into CPP2 due to a lack of circulating crystallization inhibitors.
2 – In vitro studies suggest that CPP2 and Pi induce calcification in VSMCs and stimulate expression of pro-calcification genes such as RUNX2, ALP (ALPL) and osterix (SP7). Simultaneously, contractility genes such as transgelin (SM22a) diminish.
3 – Combined, this cascade results in VSMC transdifferentiation and calcification and loss of VSMC function and contractility.
4 – The resulting calcified VSMCs amplify the calcification process by shedding Ca- loaded exosomes that are engulfed by neighbouring VSMCs.
the pathogenesis of vascular calcification has 2 main mechanisms:
1- the transdifferentiation of vascular smooth muscle into osteoblast like cells
2- formation of secondary calciprotein particles
The medial calcification in Ckd occur due to the VSMC transdifferentiate to osteoblast like cell with expression of gene like RUNX, BMP, osteopontin, osx and alkaline phosphotase, these will cause increase of extracellular matrix and release of Ca from exosome, these initiated by increase phosphorus.
As phosphorous increase with Calcium and decrease inhibitor (fetuin A, albumin, osteopontin) so they combined to form amorphous soluble Ca-ph particle or primary CCP which mature to form secondary CCP which is form needle shape calcification in the medial layer of vessels which can be prevented by high Mg because once it occur cannot inhibit the process. The maturation time from CCP1 to CCP2 called T50 which is the measure of of calcification propensity in indiv
Recent studies have shown that magnesium can inhibit the maturation of calciprotein particles (CPPs), which are implicated in the development of vascular calcification.
Additionally, magnesium can protect the mineral-buffering system that is overwhelmed by phosphate in CKD, thereby reducing the risk of mineral deposition in the vasculature.
Magnesium also appears to play a role in preventing vascular smooth muscle cells (VSMCs) from transdifferentiating into an osteoblast-like phenotype, thereby preventing the expression of osteogenic genes and the production of calcification-promoting substances.
A clinical perspective of modifying Mg balance
Recently as CPP formation depends on Ca, Pi, and futin-A concentration, CPP may explain Pi-stimulated vascular calcification in CKD and thus mediate Pi-induce VSMC calcification, so;
Magnesium can prevent CPP2 formation by reducing Pi effect, and this supports the need to quantify the mineral-buffering capacity in CKD patients to withstand vascular calcification, and this may open a promising window toward a potential treatment target for vascular calcification.
Meanwhile, measures have to be taken to reduce this risk;
In view of recent studies showing that CPP2 induces VSMC transdifferentiation and vascular calcification, the hypothesis indicating a prominent role for Mg2+ on intracellular pro-calcifying pathways may be insufficient.
CPP2 may develop before VSMCs become osteoblast-like. Mg2+ inhibition of CPP2 maturation will diminish osteogenic gene expression and keep VSMCs contractile.
Recent studies demonstrate that magnesium suppresses calciprotein particle formation, which may explain its anti-calcification effects. Magnesium protects the calcification milieu and restores the mineral-buffering system that phosphate overwhelms in CKD patients.
Magnesium is a regulator of calciprotein particle maturation and extracellular matrix mineralization, making it a viable new therapeutic strategy for treating CKD vascular calcification.
the evolution and the recent explanation and understanding the pathogenesis of vascular calcification in CKD and in dialysis pt.
and more study and research about new non traditional risk factors as role of Mg. Calcipotriene particle maturation , Pi, FGF 23 , …..other
will change our perspective in the pathogenesis of VC and promising novel treatment and practical guide lines to decrease risk VC and mortality
How has our perspective of VC pathogenesis changed in recent years?
Three steps:
How does it happened ?