Potential of animal-welfare compliant and sustainably sourced serum from pig slaughter blood

The animal product most used as a stimulatory additive for cell cultivation is still fetal bovine serum (FBS). Besides the ethical concerns regarding serum collection, the main problems of FBS are batch-to-batch variability and the resulting risk of lower reproducibility, the differences between species, the presence of undefined/unknown components, and the risk of contamination. In contrast, pig blood, which is a by-product of slaughter, is a sufficiently available and sustainable resource with a high degree of standardization in terms of donor age, weight, and genetics. The variations in preparations from pig slaughter blood seem to be comparatively low, and consequently, batch effects might be much smaller, suggesting that the reproducibility of the research data obtained may be increased. Our pilot study aimed to investigate, as a proof of concept, whether adult human and porcine stem cells of different tissue origins proliferate and differentiate adequately when FBS is completely or partially replaced by porcine serum (PS). We could show that the human and porcine stem cells were vital and proliferated under partial and full PS supplementation. Furthermore, using PS, the two cell types studied showed tissue-specific differentiation (i.e., lipid vacuoles as a sign of adipogenic or myotubes as a sign of myogenic differentiation). In conclusion, the pig slaughter blood–derived serum has promising potential to be a replacement for FBS in adult stem cell cultures. Therefore, it could serve as a basis for the development of new cell culture supplements. Supplementary Information The online version contains supplementary material available at 10.1007/s00441-024-03904-8.


Human subjects
For the purpose of this study, all human donors gave written informed consent for tissue donation.In total, material from five patients (female) who had undergone tumescent liposuction was used.The mean age of the donors was 46.2 ± 3.1 years (between 43 and 50 years) with an average body mass index of 30.4 ± 3.0 kg/m 2 .

Animals
The animals were obtained from the Experimental Animal Facility Pig of the Research Institute for Farm Animal Biology (FBN), Dummerstorf, Germany (70 days-old pigs) or from the Testing station of the Hybrid Pig Breeding Association North/East MV (fattening pigs) and were not part of animal experiments.Animal husbandry and slaughter followed the guidelines set by the Animal Care Committee the State of Mecklenburg-Western Pomerania, Germany, based on the German Law for Animal Protection.
For serum collection, ten female fattening pigs (171 ± 15 days of age) with a body weight of 124 ± 5 kg were killed using exsanguination after electro stunning.
For muscle cell culture, five female 70-day-old pigs with a body weight of 19.4 ± 1.1 kg were killed at the FBN Experimental abattoir using exsanguination after electro stunning.Tissue of the M. rhomboideus was dissected and washed in enriched phosphate buffered saline until the isolation procedure.

Table 3 :
Serum-dependent normalized cell indices (nCI) and slopes of human adMSC and porcine musSC over a period of 72 hours Impedance measurements (xCELLigence, ACEA Biosciences Inc) were recorded every 30 min for up to 96 hours and results were expressed as nCI.The nCI and slope values provided by the system (Software RTCA, Version 1.2.1.)between24 and 96 hours were used for our statistical analyses.Numerical data are presented as means with SEM.When the Shapiro-Wilk test indicated a non-Gaussian distribution, statistical analysis was performed using the Kruskal-Wallis test, followed by a Dunn multiple comparison test with *P < 0.05 compared to the control (Ctrl.).For data with a Gaussian distribution, an ANOVA test (Ordinary one-way analysis of variance) was performed, followed by a Dunnett post hoc test (multiple comparison) with *P < 0.05 compared to control.The experiments were performed with three independent adMSC or musSC preparations and three technical replicates.FBS: fetal bovine serum; PS: porcine serum