Good Manufacturing Practice-Compliant Cryopreserved and Thawed Native Adipose Tissue Ready for Fat Grafting.

Background: As adipose tissue-derived mesenchymal stem cells are becoming the tool of choice for many clinical applications; standardized cryopreservation protocols are necessary to deliver high-quality samples. For this purpose, the cryopreservation and thawing of native adipose tissue under GMP conditions could represent an extremely useful and powerful tool for the direct reinfusion of the tissue, and consequently, of its stromal vascular fraction. Methods: In this study, 19 samples of adipose tissue were cryopreserved and characterized before and after storage in liquid nitrogen vapors. Of these 19 samples, 14 were processed in research and 5 in a GMP-compliant environment. Storage with and without cryopreservation medium was also evaluated. After one week to three months of storage, samples were thawed, washed, enzymatically digested, and characterized with flow cytometry. Results: The results show that there is a loss of nearly 50% of total nucleated cells during the cryopreservation/thawing process. Non-GMP and GMP samples are comparable for all parameters analyzed. This study also allowed us to exclude the cryopreservation of adipose tissue without any cryopreservation medium. Conclusions: The data shown in this work are consistent with the idea that native adipose tissue, if properly processed and controlled, could be a useful source of cells for regenerative medicine, keeping in mind that there is a clear difference in the quality between fresh and thawed samples.

Analytical investigations on elastomeric shells of new Poly Implant Prothèse (PIP) breast and from sixteen cases of surgical explantation.

In this study we have analysed the chemical composition of the extractable components from the elastomeric shells from n=27 explanted implants (n=16 patients), n=2 new PIP implants, and from explanted Eurosilicone and McGhan implants (n=2 for each brand). Empty elastomeric PIP shells (n=2) were also available for analysis. Prostheses were explanted between 2011 and 2012, after 3-11 years of implantation. Patients presented pre- and post-operation complications of different degrees. First we evaluated the micro-structural features of the shells by phase contrast microscopy, and compared the results with those obtained using attenuated total reflectance infrared spectroscopy (ATR-FT-IR), head space gas chromatography coupled to mass spectrometry (HS-GC-MS) and GC-MS in direct injection mode. The results of these studies confirm and explain the conclusion of previous international investigations that evidenced a linear decrease of rupture rate for prostheses implanted after 2006. The analytical identification of a series of UV sensitive radical photo-initiators (0.1%-0.2% w/w), belonging to the benzophenone family, in the shells of new and explanted prostheses produced after 2006 (associated with a strong smell absent in those produced earlier), in parallel to (ii) the removal of the anti bleed barrier from their manufacturing procedure (introduced after 2001), strongly suggested that from 2007, the PIP company used a polymerisation process different from that declared, probably as a consequence of the first reports of rupture problems. In addition, the HS-GC-MS experiments demonstrated that at body temperature the diffusivity of low molecular weight (LMW) silicones present in the PIP filler silicone is significantly higher than that of LMW silicones in an approved implant.

Chemical and biochemical composition of late periprosthetic fluids from women after explantation of ruptured Poly Implant Prothèse (PIP) breast prostheses.

In this study we have analysed the chemical composition of the silicone extracted from two explanted intact PIP breast prostheses and of breast late periprosthetic fluid (LPF) samples from n=4 patients with ruptured PIP implants. The results obtained by ATR-FT-IR spectroscopy, GC-MS, reverse phase HPLC-UV/DAD, SEC-UV/DAD and contrast phase microscopy demonstrated for the first time that the cloudy, viscous LPF found in the breast of women carriers of PIP implants is a multiphasic amphiphilic silicone/serum microemulsion that can migrate through the body (via lymphatic system) to accumulate first in the lymph nodes. The GC-MS and ATR-FT-IR data indicate that low and high molecular weight silicones (chemical markers: D4-D9, L8) can penetrate the elastomeric shell and periprosthetic capsule and enter the breast tissue via the periprosthetic fluid. Also serum influx from breast tissue into via periprosthetic fluid through the capsule and shell into implant filler silicone (chemical markers: cholesterol by GC-MS, and uric acid, globulins and albumin by HPLC-DAD). To the best of our knowledge, the profile of the major constituents in PIP-induced LPF have been unequivocally characterised for the first time in this work. Further studies will be needed to evaluate the biological consequences of the current results. The potential toxicological implications of the results are discussed in the light of the current literature on the health effects of PIP implants.

Chemical and physicochemical properties of the high cohesive silicone gel from Poly Implant Prothèse (PIP) breast prostheses after explantation: a preliminary, comparative analytical investigation.

Aim of this work was to gain a deeper insight into the analytical profile of the macromolecular and LMW fractions of polymeric silicones present in breast implants. The study was conducted on silicone gel samples from (i) breast prostheses (Poly Implant Prothèse, PIP) explanted from a patient that needed their therapeutical removal, (ii) from a virgin Mc Ghan 410 MX prosthesis and (iii) from a sample of technical-grade non-cohesive silicone. The gels were analysed using rheological techniques, attenuated total reflectance infrared spectroscopy (ATR-FT-IR), nuclear magnetic resonance ((1)H NMR), gas chromatography coupled to mass spectrometry (GC-MS) and flow injection electrospray mass spectrometry (FI-ESI-MS). Our results demonstrate that, compared to the virgin McGhan gel, the silicone present the PIP prostheses lacks a significant part of the cross-linking sites necessary for the high-cohesive properties of the gel, significant amounts of cholesterol have been absorbed from the breast tissue by the silicone material, demonstrating the lack of impermeability of its elastomer shell. The potential implications and consequences of these analytical results are discussed.