TGFβ activity released from platelet-rich fibrin adsorbs to titanium surface and collagen membranes

Summa, Francesca Di; Kargarpour, Zahra; Nasirzade, Jila; Stähli, Alexandra; Mitulović, Goran; Panić-Janković, Tanja; Koller, Veronika; Kaltenbach, Cosima; Müller, Heinz; Panahipour, Layla; Gruber, Reinhard; Strauß, Franz Josef

doi:10.1038/s41598-020-67167-3

Cited by 29 publications

(57 citation statements)

References 58 publications

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“…Given the proposed functions of DCN and BGN, SLRPs may be involved in the control of TGF-β activity and collagen fibril formation in the peritoneum [ 24 ]. Our previous in vitro data support the binding capacity of collagen membranes for TGF-β [ 41 , 42 ]. Thus, the identification of various leucine rich proteins in both the membrane and the matrix reflects the composition of the original tissue and may translate into a biological activity in the biomaterial.…”

Section: Discussionsupporting

confidence: 76%

Proteomic Analysis of Porcine-Derived Collagen Membrane and Matrix

et al. 2020

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Collagen membranes and matrices being widely used in guided bone regeneration and soft tissue augmentation have characteristic properties based on their composition. The respective proteomic signatures have not been identified. Here, we performed a high-resolution shotgun proteomic analysis on two porcine collagen-based biomaterials designed for guided bone regeneration and soft tissue augmentation. Three lots each of a porcine-derived collagen membrane and a matrix derived from peritoneum and/or skin were digested and separated by nano-reverse-phase high-performance liquid chromatography. The peptides were subjected to mass spectrometric detection and analysis. A total of 37 proteins identified by two peptides were present in all collagen membranes and matrices, with 11 and 16 proteins being exclusively present in the membrane and matrix, respectively. The common extracellular matrix proteins include fibrillar collagens (COL1A1, COL1A2, COL2A1, COL3A1, COL5A1, COL5A2, COL5A3, COL11A2), non-fibrillar collagens (COL4A2, COL6A1, COL6A2, COL6A3, COL7A1, COL16A1, COL22A1), and leucine-rich repeat proteoglycans (DCN, LUM, BGN, PRELP, OGN). The structural proteins vimentin, actin-based microfilaments (ACTB), annexins (ANXA1, ANXA5), tubulins (TUBA1B, TUBB), and histones (H2A, H2B, H4) were also identified. Examples of membrane-only proteins are COL12A1 and COL14A1, and, of matrix only proteins, elastin (ELN). The proteomic signature thus revealed the similarities between but also some individual proteins of collagen membrane and matrix.

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Section: Discussionsupporting

confidence: 76%

Proteomic Analysis of Porcine-Derived Collagen Membrane and Matrix

et al. 2020

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“…Thus, we determined whether blocking of the TGF-β receptor 1 kinase with SB431542 affects the expression of IL11. In support of the notion that IL11 is a highly sensitive TGF-β target gene [13][14][15] and even mediates the effects of TGF-β in vivo [16,17], SB431542 completely blocked the expression when activated with the supernatant but also the lysates of MG and CG2 ( Figure 5). Further support for the TGF-β activity in the supernatant of MG and CG2 comes from findings that the TGF-β neutralizing antibody reduced IL11 mRNA expression in two independent experiments from 19.0/18.3 to 3.2/1.4 (MG) and 2.0/28.3 to 0.7/0.9 (CG2).…”

Section: Tgf-β Receptor I Kinase Is Required To Increased Tgf-β Targesupporting

confidence: 61%

“…Moreover, and considering that the detection of TGF-β on the protein level not necessarily allows conclusions on the activity of the growth factor, we have performed a bioassay, refining previous observations to a functional level. This bioassay was used to report the TGF-β activity of bone conditioned medium [22], acid bone lysate [21], enamel matrix derivatives [14], and lysates of PRF membranes [13]. Surprisingly, TGF-β is not part of the proteomic signature of the MG membrane (Lee et al unpublished) with maybe attributed to the detection limit of the method; it thus seems that a bioassay with oral fibroblasts being highly sensitive to TGF-β signaling is an appropriate cellular tool to identify this growth factor activity.…”

Section: Discussionmentioning

confidence: 99%

“…The overall aim of the present research is therefore to identify the TGF-β activity that is released from a panel of collagen membranes either spontaneous or upon their digestion with collagenase. We took advantage of our established bioassays allowing us to measure the TGF-β activity that is based on the expression of at least three sensitive target genes, namely IL11, NOX4, and PRG4 that were among the most sensitive genes in gene arrays [13][14][15]. Both IL11 and NOX4 are not only target genes, as there are also critically involved in mediating downstream TGF-β effects in cardiovascular and liver fibrosis [16,17] and systemic sclerosis [18].…”

Section: Introductionmentioning

confidence: 99%

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TGF-β Activity Related to the Use of Collagen Membranes: In Vitro Bioassays

Panahipour

Kargarpour

Luza

et al. 2020

IJMS

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Collagen membranes commonly used in guided bone regeneration are supposed to actively influence tissue regeneration and are not exclusively serving as passive barriers shielding away the soft tissue. The molecular mechanisms by which collagen membranes might affect tissue regeneration might involve the activation of transforming growth factor beta (TGF-β) signaling pathways. Here, we determined the TGF-β activity of supernatants and proteolytic lysates of five commercially available collagen membranes. The expression of TGF-β target genes interleukin 11 (IL11), NADPH oxidase 4 (NOX4), and proteoglycan 4 (PRG4) was evaluated by reverse transcriptase polymerase chain reaction and IL11 immunoassay in gingival fibroblasts. TGF-β signaling activation was further assessed by blocking the TGF-β receptor I kinase, a TGF-β neutralizing antibody, and showing the nuclear localization of phosphorylated Smad3 and total Smad2/3. We could identify two collagen membranes whose supernatants and lysates caused a robust increase of TGF-β receptor I kinase-dependent expression of IL11 in gingival fibroblasts. Moreover, the supernatant of a particular one membrane caused the nuclear localization of phosphorylated Smad3 and Smad2/3 in the fibroblasts. These results strengthen the evidence that some collagen membranes possess an intrinsic TGF-β activity that might actively influence the process of guided bone regeneration.

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“…Heating, however, not only inactivates plasminogen and coagulates albumin; it might also affect the activity of the catalase [21] and growth factors intrinsic to the PRF clot. We recently performed a proteomic analysis combined with a whole genome gene array of a traditional PRF membrane and identified that TGF-β was a major growth factor with respect to its activation of genes in oral fibroblasts [22]. TGF-β activity is identified by its ability to regulate respective target genes, most notably IL11 and NOX4, that appeared in multiple independent screening approaches involving enamel matrix proteins [23], bone conditioned medium [24] and acid bone lysates [25].…”

Section: Introductionmentioning

confidence: 99%

Liquid Platelet-Rich Fibrin and Heat-Coagulated Albumin Gel: Bioassays for TGF-β Activity

et al. 2020

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Liquid platelet-rich fibrin (PRF) can be prepared by high centrifugation forces separating the blood into a platelet-poor plasma (PPP) layer and a cell-rich buffy coat layer, termed concentrated PRF (C-PRF). Heating the liquid PPP was recently introduced to prepare an albumin gel (Alb-gel) that is later mixed back with the concentrated liquid C-PRF to generate Alb-PRF. PRF is a rich source of TGF-β activity; however, the overall TGF-β activity in the PPP and the impact of heating the upper plasma layer remains unknown. Here, we investigated for the first time the in vitro TGF-β activity of all fractions of Alb-PRF. We report that exposure of oral fibroblasts with lysates of PPP and the buffy coat layer, but not with heated PPP, provoked a robust increase in the TGF-β target genes interleukin 11 and NADPH oxidase 4 by RT-PCR, and for IL11 by immunoassay. Consistent with the activation of TGF-β signaling, expression changes were blocked in the presence of the TGF-β receptor type I kinase inhibitor SB431542. Immunofluorescence and Western blot further confirmed that lysates of PPP and the buffy coat layer, but not heated PPP, induced the nuclear translocation of Smad2/3 and increased phosphorylation of Smad3. The immunoassay further revealed that PPP and particularly BC are rich in active TGF-β compared to heated PPP. These results strengthen the evidence that not only the cell-rich C-PRF but also PPP comprise a TGF-β activity that is, however, heat sensitive. It thus seems relevant to mix the heated PPP with the buffy coat C-PRF layer to regain TGF-β activity, as proposed during the preparation of Alb-PRF.

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TGFβ activity released from platelet-rich fibrin adsorbs to titanium surface and collagen membranes

Cited by 29 publications

References 58 publications

Proteomic Analysis of Porcine-Derived Collagen Membrane and Matrix

Proteomic Analysis of Porcine-Derived Collagen Membrane and Matrix

TGF-β Activity Related to the Use of Collagen Membranes: In Vitro Bioassays

Liquid Platelet-Rich Fibrin and Heat-Coagulated Albumin Gel: Bioassays for TGF-β Activity

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