Efficacy of microneedling plus human stem cell conditioned medium for skin rejuvenation: a randomized, controlled, blinded split-face study
Authors: Eo Gin Lee
Dong Hyun Kim
Sangjin Kang
Hyung-Min Chung
Hee Jung Lee
Jong-Hyuk Sung
Journal: Annals of dermatology (2014)
Discipline: CLINICAL MEDICINE
Keywords: Aging; Conditioned culture media; Embryonic stem cell; Microneedling; Rejuvenation
Affiliations: Department of Dermatology, CHA Bundang Medical Center, CHA University, Seongnam, Korea., Department of Applied Bioscience, CHA University, Pocheon, Korea., Department of Applied Bioscience, CHA University, Pocheon, Korea. ; College of Pharmacy, Yonsei University, Incheon, Korea., Stem Cell Research Laboratory, Department of Biomedical Science, CHA Stem Cell Institute, CHA University, Seoul, Korea. ; Department of Stem Cell Biology, Konkuk University School of Medicine, Seoul, Korea.

Abstract:  Background: The use of growth factors in skin rejuvenation is emerging as a novel anti-aging treatment. While the role of growth factors in wound healing is well established, their use in skin rejuvenation has only recently been to be studied and no controlled trials have been performed. Objective: We evaluated the anti-aging effects of secretory factors of endothelial precursor cells differentiated from human embryonic stem cells (hESC-EPC) in Asian skin. Methods: A total of 25 women were included in this randomized, controlled split-face study. The right and left sides of each participant's face were randomly allocated to hESC-EPC conditioned medium (CM) or saline. To enhance epidermal penetration, a 0.25-mm microneedle roller was used. Five treatment sessions were repeated at 2-week intervals. Results: Physician's global assessment of pigmentation and wrinkles after treatment revealed statistically significant effects of microneedling plus hESC-EPC CM compared to microneedling alone (p&lt;0.05). Skin measurements by Mexameter and Visiometer also revealed statistically significant effects of microneedling plus hESC-EPC CM on both pigmentation and wrinkles (p&lt;0.05). The only minimal adverse event was mild desquamation in one participant. Conclusion: Secretory factors of hESC-EPC improve the signs of skin aging and could be a potential option for skin rejuvenation. 

Retracted: No

PubPeer comments:
 From the Methods Materials section: * Study of sequence similarities were performed using the Align tool of Uniprot (uniprot.org). For NTCP, the sequence UniProtKB - Q14973 (NTCP_HUMAN) was used. For DBP-MAF, the sequence UniProtKB - P02774 (VTDB_HUMAN) was used. For HBSAG, the sequence UniProtKB - P03138 (HBSAG_HBVD3. Isolate France Tiollais 1979) was used. For alpha-N-acetylgalactosaminidase (abbreviated in NAGAB_HUMAN), the sequence UniProtKB - P17050 (NAGAB_HUMAN) was used. The sequences **TNGTKR, HKNNKS, RSYLTPGDSSSG, and QTNSPRRA** were retrieved from Jaimes et al. (10).* Then in the Results, * This concentration of negatively charged aminoacids may be involved in the binding and electrostatic neutralization of protein inserts characterized by high density of positively charges residues. In particular, it may be involved in binding the sequences **TNGTKR, HKNNKS, RSYLTPGDSSSG, and QTNSPRRA** that are characterized by a high surface concentration of positive charges interspersed with hydrophobic residues (10, 13). * Refs [10] and [13] are -  Structural modeling of 2019-novel coronavirus (nCoV) spike protein reveals a proteolytically-sensitive activation loop as a distinguishing feature compared to SARS-CoV and related SARS-like coronaviruses . -  Furin, a potential therapeutic target for COVID-19 . It is not immediately clear why the present authors decided to include **SARS-CoV-19** spike proteins in a manuscript purportedly devoted to sodium-bile acid cotransporter. A possible clue can be found in a later sentence,  *negatively charged glycosaminoglycans such as low-molecular-weight chrondroitin sulfate (14) might also serve the scope of binding and neutralizing those sequences.* Ref [14] is  [Rationale for the design of a novel tool for immunotherapy based on an emulsion of glycosaminoglycan]  , by authors Ruggiero Pacini; it is an advertisement, published in a predatory journal, for their chondroitin-based product imuno . The third author recently published [a Powerpoint presentation] , with the emphatic claim that  [We found the coronavirus vulnerability: Learn how to exploit it]  . It leads up to the claim that since chondroitin sulphate preparations inhibit HIV-1,  u2026 and [chondroitin sulphate] may work against other viruses that use positively charged molecules of their surface to attack their hosts u2026 By  other viruses , Ruggiero specified SARS-CoV-19, alluding to the discredited claim that SARS-CoV-19 was synthesised in a laboratory using segments of the HIV-1 genome. ![file] ![file] The presentation finishes by promoting the authors fermentation-based products as sources of DBF-MAF and chondroitin sulphate. Those products also feature in the text of the paper:  We demonstrated that, in addition to its presence in human serum, DBP-MAF is formed during fermentation of milk and colostrum (6) as well as during fermentation of hemp seed proteins (7). ![file] It happens that previous chondroitin-related papers by authors Pacini and Ruggiero found their way into a [promotional  Science supports Imuno! webpage set up by that product s manufacturer distributor] , and it is reasonable to surmise [that it was published with this purpose in mind] . We should at least consider the possibility that this manuscript, if it is accepted for publication in a journal, would find its way onto that webpage in turn, and be used in claims for Imuno as a cure for COVID-19. ![] , 
 I conjectured in #1 that the present manuscript is intended to prepare a foundation for a claim that the authors commercial product Imuno (an emulsion of chondroitin and vitamin D) can be repurposed as a cure for COVID-19. In fact this was foreshadowed in a [19 February blogpost] by Mimi Castellanos, a retailer of Imuno who channels communications from the last author. It is possible, then, that the manuscript is an exercise in motivated reasoning, seeking a rationale for the prediction of a curative property. *Dr. Ruggiero says, u201cI take the liberty to write that I am 99% (100% would be unrealistic) confident that imuno will help with the coronavirus for two reasons. The first and most obvious is its general effect on the immune system as described in all the papers where it has been mentioned.u201d* *u201cThe second is related to the specific anti-viral properties of chondroitin sulfate that are further amplified by the molecular arrangement of imuno. As you may know by now, the coronavirus responsible for the current epidemic attacks human cells because it has spikes from HIV that no one knows, at the present, whether they were intentionally inserted or are result of a casual viral recombination. (highly improbable but not impossible)u201d* * Needless to say, the chondroitin sulfate in imuno is much more potent bioavailable that the molecules used in those studies for at least two reasons: it is homogeneous and low-molecular-weight; it is arranged in a supramolecular structure that makes it more bioavailable and efficient in binding to the HIV spikes. * * Because of these evidences, I am 99% sure that imuno will work against coronavirus. * , 
 We read that one of the experiments was performed independently by R.E.D. Labs, of Belgium: * Study of in vitro interaction between DBP-MAF or the product of microbial fermentation of milk and colostrum mentioned above, here abbreviated in PMF (Product of Microbial Fermentation), and NAGAB_HUMAN was was performed by R.E.D. Laboratories (Zellik, Belgium) where NAGAB_HUMAN and DBP-MAF had been purified [...] [Authors MR and SP] had no prior knowledge of the results of the experiments with NAGAB_HUMAN that were independently performed by R.E.D. Laboratories (Zellik, Belgium). * Now another of author MR s projects is a probiotic food supplement, Immortalis Klotho . The marketing for this product promises to [prolong the consumer s life indefinitely](http: archive.is tL1b7) - providing literal, unambiguous immortality. The price of Immortalis is really quite extreme, as if to elevate it to the class of Veblen goods which are consumed as status symbols so that actual function is undesirable. The rationale for Klotho s implausible qualities involves quantum entanglement and relativistic effects (due to the extreme density of chondroitin molecules) causing time dilation in the vicinity of chromosomes - see [Ruggiero (2017)] . All this would be a private matter between MR and his wealthy customers, with no bearing on the pre-print on hand, except for the involvement of R.E.D. Labs as parties or partners in the promotion of this expensive placebo. They have teamed up with the Immortalis company to provide [a panel of blood tests](http: blog.redlaboratories.be 2019 03 blood-panel-test-for-immortalis.html) for convincing customers that the product is legitimate: * R.E.D. Laboratories has created a bespoke blood panel test for Immortalis customers. It is recommended to do the labs before beginning and 6 months after beginning Immortalis Klotho Formula. The Immortalis testing panel includes 6 tests:... * The pre-print on hand places some emphasis on the arms-length independence of R.E.D. Labs and of their contribution. However, their complicity with the authors in a problematic Alt-Med operation brings that independence into question, to my mind at least. , 
  We demonstrated that, in addition to its presence in human serum, DBP-MAF is formed during fermentation of milk and colostrum (6) as well as during fermentation of hemp seed proteins (7). Ref. [6] is  6. PacinitS, Punzi T, MoruccitG, Ruggiero M. Macrophages of the mucosa- associated lymphoidttissue (MALT) as key elements of the immune response to vitamin d binding protein macrophage activating factor. It J Anat Embryol. 2011; 116(1): 136. doi: 10.13128 IJAE-10160 https: www.torrossa.com en catalog preview 2490419 . In that one-page report, the authors **assumed** that a fermentation of milk with a proprietary mixture of microbes creates DBP-MAF within the resulting Magic Yoghurt... contrary to the description here, they did not measure the protein or test for its presence. Ref [7] turns out to be a lengthy footnote rather than a citation. * The process of fermentation of hemp seed protein extract was performed using the following materials a proprietary probiotic formula containing Lactobacillus salivarius, Lactobacillus acidophilus, Lactobacillus paracasei, Lactobacillus rhamnosus, Lactococcus lactis, Bifidobacterium infantis, Bifidobacterium bifidum, Bifidobacterium lactis, Bifidobacterium longum, and proprietary cultured kefir grains whose peculiar microflora was recently described (8). The fermented hemp seed protein extract caused inhibition of HCT116 cell proliferation that was statistically significant (p 0.05). We attribute this effect to formation of bioactive peptides from edestin. Edestin is an hexameric globular protein that bears striking functional resemblance to DBP-MAF. We postulate that proteases secreted by the microbes of the probiotic formulation, together with the increase of acidity that is characteristic of fermentation, led to formation of peptides functionally very similar to DBP-MAF and, in particular, to the peptide TPTELAKLVNKRSE that was identified as the active site of DBP-MAF (9) * I can only suppose that the original intention had been to link to a paper here*, but for some reason this was not possible, so it was replaced with the footnote *summarising* the paper. The claim that the hemp protein Edestin  bears striking functional resemblance to DBP-MAF calls out for supporting evidence (although similar claims appear to be widespread in the hemp-product industry). At least its name explains why the authors are now producing and selling the hempseed-enhanced yoghurt as  Bravo-Edestiny . https: www.healthyenergetics.com collections bravo-edestiny Edestiny. * Perhaps [Ruggerio Pacini (2020)] ,  Fermentation of hemp seed proteins leads to formation of peptides that share sequence similarity with human vitamin D-binding protein - from the predatory publisher OAText. ,