The cell line used in the study is ATDC5 which is a rat cell line so human chondrocyte cells might have different results.
The mechanism of ascorbic acid-induced differentiation of ATDC5 chondrogenic cells.
"collagen II hydrogels could stimulate the differentiation of bone marrow mesenchymal stem cells in the absence of growth factors."<-Thus injecting type II collagen hydrogels into your bone marrow could make you taller.
Since the compounds of Vitamin C and the other compounds of the medium are so readily available in the body it's likely to be a problem with the stem cells themselves that's inhibiting chondrogenic differentiation.
Thus study was done on adult stem cells:
MSC proliferation and the number of colony forming units were increased by ascorbic acid, whereas dexamethasone enhanced the proportion of ALP-positive CFU and was critical for mineral deposition. Runx-2 and type I collagen gene expression decreased along with additive-induced MSC differentiation[we don't want Runx-2 and type I collagen as that is osteogenic while we want chondrogenic so we would not want Beta-glyerophosphate(dexamethasone has been found to have potential height decreasing effects)], i.e. from C1 to C4, while ALP and osteocalcin were differently regulated."
Chondrogenic markers were not measured in this study. However, using a different cell line produced very different results in regards to Vitamin C.
Here's a study that shows that age doesn't affect stem cell differentiation however:
"Expanded MSCs were released by trypsin treatment, counted and resuspended at a density of 1 × 106. The medium was changed to 500 μl DMEM with 10% FCS, 1% antibiotic mix (penicillin/streptomycin), 37.5 μg/ml (100 μmol/l) ascorbate-2 phosphate, and 10–7 mol/l dexamethasone. Pellet cultures were incubated with 10 ng/ml recombinant human transforming growth factor-β3 during chondrogenesis. After 3 weeks the samples were used."
"Pellet cultures of MSCs resulted in the formation of dense nodules consistent with chondrogenic differentiation."<-So forming a pellet culture is most important in inducing chondrogenesis. Can hydrostatic pressure like that induced by LSJL result in the MSC formation of pellet cultures?
"CD9{down in LSJL}, a mediator of tetraspanin protein-driven cellular adhesion, invasion, and fusion, was verified in mesenchymal cells during expansion "
So Vitamin C enhances both chondrogenic and osteogenic differentiation. Age does not seem to affect whether differentiation is headed towards an osteogenic or chondrogenic path but rather how the stem cells are cultured. Hydrostatic pressure may change the culture within the bone marrow to be more chondrogenic.
Ascorbate-enhanced chondrogenesis of ATDC5 cells.
"The ATDC5 cell line exhibits the multistep chondrogenic differentiation observed during endochondral bone formation. However, it takes up to two months to complete the process of cell expansion, insulin addition to promote differentiation and further changes in culture conditions effectively to induce hypertrophy. By adding ascorbate, the prechondrogenic proliferation phase was shortened from 21 to 7 days, with production of cartilaginous nodules during the chondrogenic phase, after insulin addition, that were greater in number and larger in size. [There was] much greater matrix elaboration and the mRNA expression of sox9, aggrecan and collagen type II were all significantly increased earlier and to a much higher degree when compared with controls. [Hypertrophic indicators] Col10a1, Runx2 and Mmp13 were all induced within 7-10 days. addition of ascorbate to ATDC5 cultures shortened the prechondrogenic proliferation phase, produced earlier chondrogenic differentiation, heightened gene expression and robust hypertrophic differentiation."
Increasing serum levels of Vitamin C, proline, and lysine may increase height but there may be negative feed mechanisms and points of diminishing returns.
Effects of both vitamin C and mechanical stimulation on improving the mechanical characteristics of regenerated cartilage.
"[We studied] vitamin C (VC) and mechanical stimulation on development of the extracellular matrix (ECM) and improvement in mechanical properties of a chondrocyte-agarose construct in a regenerating tissue disease model of hyaline cartilage. We used primary bovine chondrocytes and two types of VC, ascorbic acid (AsA) as an acidic form and ascorbic acid 2-phosphate (A2P) as a non-acidic form, and applied uniaxial compressive strain to the tissue model using a purpose-built bioreactor. When added to the medium in free-swelling culture conditions, A2P downregulated development of ECM and suppressed improvement of the tangent modulus more than AsA. Application of mechanical stimulation to the construct both increased the tangent modulus more than the free-swelling group containing A2P and enhanced the ECM network of inner tissue to levels nearly as high as the free-swelling group containing AsA. Mechanical stimulation and strain appears to enhance the supply of nutrients and improve the synthesis of ECM via mechanotransduction pathways of chondrocytes. Mechanical stimulation is necessary for homogenous development of ECM in a cell-associated construct with a view to implantation of a large-sized articular cartilage defect."
"Even though A2P has no physiological activity, it can nevertheless reproduce the effect of VC activity after dephosphorylation by an alkaline phosphatase (ALP). Dephosphorylated A2P, or AsA, penetrates chondrocytes through a VC transporter, chiefly the sodium-dependent VC transporter 2 (SVCT2), and supports collagen synthesis as a cofactor in the rough endoplasmic reticulum. In free-swelling culture conditions, we only observed a small number of collagen molecules distributed in the construct. We also observed that the tangent moduli of both A2P dose groups were lower than the moduli of AsA(2.2). We must also consider the reaction rates of both dephosphorylation of A2P by ALP and the transport of AsA into the cytosol via SVCT2."
Effect of different ascorbate supplementations on in vitro cartilage formation in porcine high-density pellet cultures.
"Most strategies [for articular cartilage regeneration] require ascorbate supplementation to promote matrix formation by isolated chondrocytes. We evaluate ascorbate forms and concentrations on in vitro cartilage formation in porcine chondrocyte high-density pellet cultures. l-ascorbate[AsA], sodium l-ascorbate[NaAs], and l-ascorbate-2-phosphate[AsAP] were administered in 100 microM, 200 microM, and 400 microM in the culture medium over 16 days. Pellet thickness increased independently from the supplemented ascorbate form and concentration. Hydroxyproline content increased as well, but here, medium concentration of AsAP and low concentration of AsA showed a more pronounced effect. Proteoglycan and collagen formation could be proven in all supplemented cultures. Non-supplemented cultures, however, showed no stable matrix formation at all. Effects on the gene expression pattern of cartilage marker genes (type I and type II collagen, aggrecan, and cartilage oligomeric matrix protein (COMP)) were studied compared to non-supplemented control cultures. Expression level of cartilage marker genes was elevated in all cultures showing that dedifferentiation of chondrocytes could be prevented. All supplementations caused a similar effect except for low concentration of AsA, which resulted in an even higher expression level of all marker genes. We could not detect a pronounced difference between ascorbate and its derivates as well as between the different concentrations."
"Ascorbate has a transcriptional effect on cartilage formation promoting type II collagen and prolyl-4-hydroxylase gene expression. At the post-transcriptional level, ascorbate is responsible for the reduction of iron to its ferrous state. This reaction is important for the hydroxylation of proline and lysine. Latter hydroxy amino acids are specific for type II collagens. Their non-covalent bonds support collagen triple helix and fibril formation. Collagen secretion from the cells is promoted by ascorbate-dependent hydroxylation within the endoplasmatic reticulum"
"Standard concentration of [Vitamin C is] about 280 μM"
"NaAs has the advantage of being non-acidic, while AsA is acidic in aqueous solutions such as culture medium. AsAP is not only non-acidic, but also characterized by a longer half-life in aqueous solutions compared to the other derivatives. It is internalized, dephosphorylated, and highly concentrated in the aqueous phase of the cells. AsA, NaAs and AsAP were administered in 100 μM, 200 μM, and 400 μM over 16 days. "
"The best results of 5 μg/mg were achieved by supplementation with 100 μM of AsA"
have you heard of the telomere activator peptide epitalon?
ReplyDeleteit increases melatonin levels, Epitalon did not influence food consumption, body weight or mean life span of mice. However, it slowed down the age-related switching-off of estrous function and decreased the frequency of chromosome aberrations in bone marrow cells (by 17.1%).