scholarly journals Human Adipose-derived Mesenchymal Stromal Cells Exhibit High HLA-DR Levels and Altered Cellular Characteristics under a Xeno-free and Serum-free Condition

2021 ◽  
Author(s):  
Phuong TM Dam ◽  
Van T. Hoang ◽  
Hue Thi Hong Bui ◽  
Le Minh Hang ◽  
Duc M. Hoang ◽  
...  
Keyword(s):  
Cultured Cells ◽  
Chromosomal Abnormalities ◽  
Human Leukocyte ◽  
Normal Karyotype ◽  
Clinical Grade ◽  
Serum Free ◽  
Surface Marker Expression ◽  
Hla Dr ◽  
Expansion Media

Abstract Background: We have observed an increased expression of negative markers in some clinical-grade, xeno- and serum-free cultured adipose-derived mesenchymal stem/stromal cell (ADMSC) samples. It gave rise to concern that xeno- and serum-free conditions might have unexpected effects on human ADMSCs. This study aims to test this hypothesis for two xeno- and serum-free media, PowerStem MSC1 media (PS) and StemMACS MSC Expansion Media (SM), that support the in vitro expansion of ADMSCs.Methods: We investigated the expression of negative markers in 42 clinical-grade ADMSC samples expanded in PS. Next, we cultured ADMSCs from seven donors in PS and SM and examined their growth and colony-forming ability, surface marker expression, differentiation, cell cycle and senescence, as well as genetic stability of two passages representing an early and late passage for therapeutic MSCs.Results: 15 of 42 clinical-grade PS-expanded ADMSC samples showed an increased expression of negative markers ranging from 2.73% to 34.24%, which positively correlated with the age of donors. This rise of negative markers was related to an upregulation of Human Leukocyte Antigen – DR (HLA-DR). In addition, the PS-cultured cells presented decreased growth ability, lower frequencies of cells in S/G2/M phases, and increased ß-galactosidase activity in passage 7 suggesting their senescent feature compared to those grown in SM. Although MSCs of both PS and SM cultures were capable of multilineage differentiation, the PS-cultured cells demonstrated chromosomal abnormalities in passage 7 compared to the normal karyotype of their SM counterparts.Conclusions: These findings suggest that the SM media is more suitable for the expansion of therapeutic ADMSCs than PS. The study also hints a change of ADMSC features at more advanced passages and with increased donor’s age. Thus, it emphasizes the necessity to cover these aspects in the quality control of therapeutic MSC products.

scholarly journals Human Adipose-Derived Mesenchymal Stromal Cells Exhibit High HLA-DR Levels and Altered Cellular Characteristics under a Xeno-free and Serum-free Condition

2021 ◽  
Author(s):  
Phuong T. M. Dam ◽  
Van T. Hoang ◽  
Hue Thi Hong Bui ◽  
Le Minh Hang ◽  
Duc M. Hoang ◽  
...  
Keyword(s):  
Cultured Cells ◽  
Chromosomal Abnormalities ◽  
Human Leukocyte ◽  
Normal Karyotype ◽  
Clinical Grade ◽  
Serum Free ◽  
Surface Marker Expression ◽  
Hla Dr ◽  
Expansion Media

Abstract Background We have observed an increased expression of negative markers in some clinical-grade, xeno- and serum-free cultured adipose-derived mesenchymal stem/stromal cell (ADMSC) samples. It gave rise to concern that xeno- and serum-free conditions might have unexpected effects on human ADMSCs. This study aims to test this hypothesis for two xeno- and serum-free media, PowerStem MSC1 media (PS) and StemMACS MSC Expansion Media (SM), that support the in vitro expansion of ADMSCs. Methods We investigated the expression of negative markers in 42 clinical-grade ADMSC samples expanded in PS. Next, we cultured ADMSCs from seven donors in PS and SM and examined their growth and colony-forming ability, surface marker expression, differentiation, cell cycle and senescence, as well as genetic stability of two passages representing an early and late passage for therapeutic MSCs. Results 15 of 42 clinical-grade PS-expanded ADMSC samples showed an increased expression of negative markers ranging from 2.73% to 34.24%, which positively correlated with the age of donors. This rise of negative markers was related to an upregulation of Human Leukocyte Antigen – DR (HLA-DR). In addition, the PS-cultured cells presented decreased growth ability, lower frequencies of cells in S/G2/M phases, and increased ß-galactosidase activity in passage 7 suggesting their senescent feature compared to those grown in SM. Although MSCs of both PS and SM cultures were capable of multilineage differentiation, the PS-cultured cells demonstrated chromosomal abnormalities in passage 7 compared to the normal karyotype of their SM counterparts. Conclusions These findings suggest that the SM media is more suitable for the expansion of therapeutic ADMSCs than PS. The study also hints a change of ADMSC features at more advanced passages and with increased donor’s age. Thus, it emphasizes the necessity to cover these aspects in the quality control of therapeutic MSC products. Graphical abstract

scholarly journals Cytokine influence on killing of fresh chronic lymphocytic leukemia cells by human leukocytes

Blood ◽  
1991 ◽  
Vol 77 (12) ◽  
pp. 2707-2715 ◽  
Author(s):  
D Cemerlic ◽  
B Dadey ◽  
T Han ◽  
L Vaickus
Keyword(s):  
Chronic Lymphocytic Leukemia ◽  
Mononuclear Cells ◽  
Protein A ◽  
Human Leukocyte ◽  
Lymphocytic Leukemia ◽  
Target Antigen ◽  
Peripheral Blood Mononuclear ◽  
Ifn Gamma ◽  
Hla Dr

Abstract The feasibility of combining the Lym-1 monoclonal antibody (MoAb) with interferon-gamma (IFN-gamma) in the treatment of chronic lymphocytic leukemia (CLL) was evaluated. We used an in vitro tumor lysis model that incorporated fresh CLL cells from 21 different patients as targets for two distinct normal human leukocyte effector subsets, neutrophils, and peripheral blood mononuclear cells (PBMCs). Lym-1 antigen (Lym-1- Ag) expression varied greatly and did not correlate with the expression of other CLL-associated antigens such as CD5, CD19, or HLA-DR. CLL cells were not lysed by neutrophils alone or with IFN-gamma in the absence of Lym-1. Neutrophil Lym-1-dependent cytotoxicity (ADCC) in the absence of IFN-gamma was weak and inconsistent. IFN-gamma exposure induced MoAb-dependent lysis of 80% of 21 CLL targets and resulted in an eightfold augmentation of neutrophil ADCC against the remainder. Cytotoxicity correlated directly and positively with Lym-1-Ag expression. Confirmation of the need for interaction between neutrophil IgG Fc receptors (Fc gamma Rs) and the Fc portion of the Lym-1 MoAb was obtained by demonstrating that purified Staphylococcus aureus Protein A (SpA) inhibited ADCC. IFN-gamma exposure caused no consistent alternations in Lym-1-Ag expression on CLL cells so that target antigen upregulation was unlikely to account for augmentation of neutrophil ADCC. PBMCs alone, exposed to interkeukin-2 (IL-2) or IFN-gamma, or with Lym-1 in the presence or absence of IL-2 or IFN-gamma were unable to lyse CLL targets. PBMCs were able to kill Raji Burkitt lymphoma cells in conjunction with Lym-1, so their ability to interact with Lym- 1-coated targets and their lytic functions appeared intact. These results emphasize the importance of examining fresh tumor cells with different leukocyte effector subsets before designing a clinical trial that combines a therapeutic MoAb with a cytokine.

scholarly journals Influence of glutamine on the phenotype and function of human monocytes

Blood ◽  
1995 ◽  
Vol 86 (4) ◽  
pp. 1564-1569 ◽  
Author(s):  
A Spittler ◽  
S Winkler ◽  
P Gotzinger ◽  
R Oehler ◽  
M Willheim ◽  
...  
Keyword(s):  
In Vitro Study ◽  
Antigen Expression ◽  
Intercellular Adhesion Molecule ◽  
Intercellular Adhesion Molecule 1 ◽  
Post Injury ◽  
Surface Marker Expression ◽  
Hla Dr ◽  
Healthy Donors ◽  
And Function

Reduced concentrations of glutamine (GLN) in plasma and skeletal muscle, defective host defense systems, and a diminished expression of the HLA-DR antigen on monocytes are important diagnostic parameters for late post-injury sepsis. In this in vitro study, we investigated whether blood monocyte-derived macrophage antigen expression and function from healthy donors is influenced by GLN. Lowering the GLN concentration in culture medium from 2 mmol/L to 200 mumol/L reduced the expression of HLA-DR by 40% (P < .001) on monocyte-derived macrophages, and decreased tetanus toxoid-induced antigen presentation. In addition, low GLN levels downregulated the expression of intercellular adhesion molecule-1 (ICAM-1/CD54), Fc receptor for IgG (Fc gamma RI/CD64), and complement receptors type 3 (CR3; CD11b/CD18) and type 4 (CR4; CD11c/CD18). A correlation was found between the phagocytosis of IgG-sensitized ox erythrocytes or opsonized Escherichia coli and the decreased expression of Fc gamma RI and CR3. Monocyte expression of CD14, CD71, and Fc gamma RIII/CD16 and capacity to phagocytose latex beads were not affected by altering the level of GLN. Depletion of GLN was associated with a significant reduction in cellular adenosine triphosphate (ATP), which may have influenced cell surface marker expression and phagocytosis. It remains to be seen whether these in vitro findings are of clinical significance in the treatment of sepsis.

Immunophenotyping of Nonneoplastic and Neoplastic Histiocytes in Cats and Characterization of a Novel Cell Line Derived From Feline Progressive Histiocytosis

2020 ◽  
Vol 57 (6) ◽  
pp. 758-773
Author(s):  
Miyuki Hirabayashi ◽  
James K. Chambers ◽  
Ayumi Sumi ◽  
Kei Harada ◽  
Makoto Haritani ◽  
...  
Keyword(s):  
Cell Line ◽  
Cultured Cells ◽  
Survival Times ◽  
Neoplastic Cells ◽  
Hla Dr ◽  
Formalin Fixed Paraffin ◽  
Formalin Fixed Paraffin Embedded ◽  
E Cadherin

Histiocytic proliferative diseases are rare in cats, and their pathogenesis is poorly understood. In the present study, 25 cases of histiocytic sarcoma (HS) and 6 of feline progressive histiocytosis (FPH) were examined, and survival times were recorded in 19 cases. The immunophenotypes of tumor cells in these cases as well as of nonneoplastic feline histiocytes were characterized using formalin-fixed, paraffin-embedded tissues. An FPH cell line (AS-FPH01) and xenotransplant mouse model of FPH were also established. The median survival time of HS (150 days) was significantly shorter than that of FPH (470 days). Immunohistochemically, nonneoplastic histiocytes were immunopositive for various combinations of Iba-1, HLA-DR, E-cadherin, CD204, CD163, CD208, and MAC387. By immunohistochemistry, dermal interstitial dendritic cells (iDCs) and macrophages were CD204+/E-cadherin−, while epidermal Langerhans cells (LCs) were CD204−/E-cadherin+. Neoplastic cells of 4 FPH and 18 HS were CD204+/E-cadherin− (iDC/macrophage immunophenotype), while 2 FPH and 2 HS were CD204−/E-cadherin+ (LC immunophenotype), and 5 HS were CD204+/E-cadherin+ (LC-like cell immunophenotype). Furthermore, immunohistochemical and western blot analyses of AS-FPH01 cells derived from E-cadherin-negative FPH revealed that cultured cells were immunopositive for both CD204 and E-cadherin in vitro and in vivo. These results indicate that the neoplastic cells of feline HS and FPH were variably positive for iDC/macrophage and LC markers, and their immunophenotype changed in different microenvironments. The novel cell line established in the present study may serve as an experimental model of FPH that will enable further molecular and therapeutic studies on this disease.

scholarly journals Impact of HLA-DR Antigen Binding Cleft Rigidity on T Cell Recognition

2020 ◽  
Vol 21 (19) ◽  
pp. 7081
Author(s):  
Christopher Szeto ◽  
Joseph I. Bloom ◽  
Hannah Sloane ◽  
Christian A. Lobos ◽  
James Fodor ◽  
...  
Keyword(s):  
T Cell ◽  
Protein Stability ◽  
T Cell Activation ◽  
Cell Activation ◽  
Human Leukocyte ◽  
Antigen Binding ◽  
Hla Class I Molecules ◽  
Hla Dr ◽  
Binding Cleft

The interaction between T cell receptor (TCR) and peptide (p)-Human Leukocyte Antigen (HLA) complexes is the critical first step in determining T cell responses. X-ray crystallographic studies of pHLA in TCR-bound and free states provide a structural perspective that can help understand T cell activation. These structures represent a static “snapshot”, yet the nature of pHLAs and their interactions with TCRs are highly dynamic. This has been demonstrated for HLA class I molecules with in silico techniques showing that some interactions, thought to stabilise pHLA-I, are only transient and prone to high flexibility. Here, we investigated the dynamics of HLA class II molecules by focusing on three allomorphs (HLA-DR1, -DR11 and -DR15) that are able to present the same epitope and activate CD4+ T cells. A single TCR (F24) has been shown to recognise all three HLA-DR molecules, albeit with different affinities. Using molecular dynamics and crystallographic ensemble refinement, we investigate the molecular basis of these different affinities and uncover hidden roles for HLA polymorphic residues. These polymorphisms were responsible for the widening of the antigen binding cleft and disruption of pHLA-TCR interactions, underpinning the hierarchy of F24 TCR binding affinity, and ultimately T cell activation. We expanded this approach to all available pHLA-DR structures and discovered that all HLA-DR molecules were inherently rigid. Together with in vitro protein stability and peptide affinity measurements, our results suggest that HLA-DR1 possesses inherently high protein stability, and low HLA-DM susceptibility.

scholarly journals Organic dust exposure alters monocyte-derived dendritic cell differentiation and maturation

2009 ◽  
Vol 297 (4) ◽  
pp. L767-L776 ◽  
Author(s):  
Jill A. Poole ◽  
Geoffrey M. Thiele ◽  
Neil E. Alexis ◽  
Angela M. Burrell ◽  
Conrad Parks ◽  
...  
Keyword(s):  
Dust Exposure ◽  
Organic Dust ◽  
Blood Monocytes ◽  
Monocyte Differentiation ◽  
Phagocytic Ability ◽  
Dendritic Cell Differentiation ◽  
Surface Marker Expression ◽  
Hla Dr ◽  
Macrophage Colony

Organic dust exposure in agricultural animal environments results in airway diseases. Dendritic cells (DCs) orchestrate inflammatory immune response in the airways, but little is known about how organic dust affects differentiation and maturation of monocyte-derived immature and mature DCs (iDCs, mDCs). Peripheral blood monocytes were differentiated in vitro into iDCs with granulocyte-macrophage colony stimulating factor + IL-4 (6 days) with and without swine facility organic dust extract (ODE, 0.1%). Unlike control iDCs, ODE-conditioned iDCs maintained key monocyte properties (increased mCD14, increased phagocytic ability) while expressing DC features [increased mCD83, HLA-DR, CD80, CD86, diminished cytokine (TNF-α, IL-6) responsiveness]. At day 6, iDCs were cultured for an additional 48 h ( days 7 and 8) with lipopolysaccharide (LPS) to induce mDCs. ODE-conditioned mDCs maintained high expression of mCD14+ and elevated phagocytosis while their DC features weakened as evidenced by decreased CD11c, CD83, HLA-DR, CD86, and CCR7 expression and reduced lymphocyte-stimulating capacity. Similar results were observed when monocytes were exposed to ODE for only the first 48 h and with ODE depleted of endotoxin. Control iDCs exposed to ODE during the final 2 days of iDC maturation ( days 7 and 8) did not differ from control (no ODE) iDCs in surface marker expression and phagocytic ability, but exhibited enhanced lymphocyte-stimulating capacity. Dust exposure alters monocyte differentiation to iDCs and prevents maturation of iDC to mDCs. The first 48 h of monocyte differentiation appears to be the susceptible period to exposure. Environmental exposures present during early monocyte differentiation may impact the critical balance of DCs in the lung.

Cytogenetics and spectral Karyotyping analysis in CLL Patients: Correlation with FISH and ZAP70 Expression

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 4844-4844
Author(s):  
Fabio Morato Oliveira ◽  
Daniel Mazza Matos ◽  
Lorena Lobo Figueiredo Pontes ◽  
Belinda Pinto Simoes ◽  
Eduardo M. Rego ◽  
...  
Keyword(s):  
Peripheral Blood ◽  
Cytogenetic Analysis ◽  
Chromosomal Abnormalities ◽  
Conflicts Of Interest ◽  
Karyotype Analysis ◽  
Calf Serum ◽  
Normal Karyotype ◽  
Fish Analysis ◽  
Interphase Cell

Abstract Abstract 4844 Cytogenetic abnormalities play an important role as prognostic factors in CLL. However, due the low mitotic index of CLL B cells in vitro, analysis of a set of subjects for the most commonly known aberrations is usually done by FISH on interphase cell. The objectives of this investigation were the use of the oligonucleotide DSP30 in combination with IL-2, as a B-cell mitogen for cytogenetic investigation in CLL and correlation among the karyotype analysis obtained (G-banding + SKY), FISH profile from unstimulated cells, ZAP70 expression and stratification status for each patient. For metaphase induction, peripheral blood mononuclear cells were cultured in RPMI 1640 medium with 20% fetal calf serum in the presence of the immunostimulatory CpG-oligonucleotide DSP30 and IL-2. Additionally, one set of cell culture was performed for each patient without any stimulant agent, for FISH analysis. The FISH panel included probes for the detection of +12, and deletions of 11q22.3 (ATM), 13q14 (D13S25 and D13S319), and 17p13 (TP53). The cut off levels for trissomy 12 (>2%), del(13q) (>2.4%), del(11q23.3) (>2.5%), del(17p13.1) (>3%) were established according to the iFISH patterns observed in a group of 4 age and sex-matched normal control peripheral blood samples studied with the same probes. Spectral karyotype analysis (SKY) was performed, according manufactures' instruction. The ZAP70 profile was obtained by flow cytometry analysis. In concordance with literature, the cut off value adopted for ZAP70 was 20%. In a group of 64 subjects studied, the cytogenetic analysis showed chromosomal aberrations in 52 patients (81.25%). The profile of abnormalities observed were del(6)(q24), +8(x2), del(11)(q13~q23), +12, +15(x2), del(12)(p13), -17, +21, +19, +18, del(13)(q31), del(14)(q24), del(17)(p13), +21, +4, +5, +11, t(1;12)(q31;p13), t(11;13)(q23;q12), t(15;18)(q11.1;q11), t(1;10)(p22;p14), t(14;22)(q32;q11), t(17;18)(q10;q10), t(9;13)(q21;q22), t(10;13)(q26;q14), t(9;12)(q12;p11), t(X;12)(p11.2;q24). Twelve patients exhibited normal karyotype (18.75%). All subjects presenting chromosomal abnormalities, by using G-banding analysis, were confirmed by SKY. In patients with normal cytogenetic, SKY analysis did not identified any criptic abnormality. Cells without any stimulant agent showed concordance with the cytogenetic profile obtained (FISH analysis). The ZAP70 expression did not show any relationship between the group of patients with chromosomal abnormalities and the group with normal karyotype. The use of the immunostimulatory oligonucleotide DSP30 in combination with IL-2 showed to be effective to induce cell cycle progression of CLL cells in vitro than others mitogens. Cytogenetic aberrations detected by G-banding in addition to FISH analysis were heterogeneous. The limited spectrum of chromosomal abnormalities seen by FISH analysis may contribute to underestimate the prognostic value, where others abnormalities may be present in patient's karyotype. These results indicate that classical cytogenetic analysis can contribute to the stratification of different subsets of CLL patients with complex karyotype associated with poor prognosis. Financial support: FAPESP (Proc. 07/52462-7). Disclosures: No relevant conflicts of interest to declare.

scholarly journals TSPO imaging-guided characterization of the immunosuppressive myeloid tumor microenvironment in patients with malignant glioma

2020 ◽  
Vol 22 (7) ◽  
pp. 1030-1043 ◽  
Author(s):  
Bastian Zinnhardt ◽  
Michael Müther ◽  
Wolfgang Roll ◽  
Philipp Backhaus ◽  
Astrid Jeibmann ◽  
...  
Keyword(s):  
Tumor Microenvironment ◽  
Suppressor Cells ◽  
Human Leukocyte ◽  
Strong Relationship ◽  
Therapy Resistance ◽  
Translocator Protein ◽  
Image Guided ◽  
Hla Dr

Abstract Background Tumor-associated microglia and macrophages (TAMs) and myeloid-derived suppressor cells (MDSCs) are potent immunosuppressors in the glioma tumor microenvironment (TME). Their infiltration is associated with tumor grade, progression, and therapy resistance. Specific tools for image-guided analysis of spatiotemporal changes in the immunosuppressive myeloid tumor compartments are missing. We aimed (i) to evaluate the role of fluorodeoxyglucose (18F)DPA-714* (translocator protein [TSPO]) PET-MRI in the assessment of the immunosuppressive TME in glioma patients, and (ii) to cross-correlate imaging findings with in-depth immunophenotyping. Methods To characterize the glioma TME, a mixed collective of 9 glioma patients underwent [18F]DPA-714-PET-MRI in addition to [18F]fluoro-ethyl-tyrosine (FET)-PET-MRI. Image-guided biopsy samples were immunophenotyped by multiparametric flow cytometry and immunohistochemistry. In vitro autoradiography was performed for image validation and assessment of tracer binding specificity. Results We found a strong relationship (r = 0.84, P = 0.009) between the [18F]DPA-714 uptake and the number and activation level of glioma-associated myeloid cells (GAMs). TSPO expression was mainly restricted to human leukocyte antigen D related–positive (HLA-DR+) activated GAMs, particularly to tumor-infiltrating HLA-DR+ MDSCs and TAMs. [18F]DPA-714–positive tissue volumes exceeded [18F]FET-positive volumes and showed a differential spatial distribution. Conclusion [18F]DPA-714-PET may be used to non-invasively image the glioma-associated immunosuppressive TME in vivo. This imaging paradigm may also help to characterize the heterogeneity of the glioma TME with respect to the degree of myeloid cell infiltration at various disease stages. [18F]DPA-714 may also facilitate the development of new image-guided therapies targeting the myeloid-derived TME.

Lipid metabolism in cultured cells. III. Cholesterol excretion process

1964 ◽  
Vol 207 (6) ◽  
pp. 1221-1225 ◽  
Author(s):  
J. Martyn Bailey
Keyword(s):  
Tissue Culture ◽  
Mammalian Cells ◽  
Cultured Cells ◽  
Rabbit Aorta ◽  
Rabbit Serum ◽  
Serum Free Medium ◽  
Free Medium ◽  
Serum Free ◽  
Cholesterol Excretion

Mammalian cells grown in tissue culture have been shown previously to take up considerable quantities of cholesterol from the growth medium. When cells grown on cholesterol-C14 supplemented medium were transferred to unlabeled medium containing serum, excretion of cholesterol into the outside medium took place. When cell cholesterol was labeled by intracellular synthesis from mevalonate-C14 precursor, it also was excreted readily into the serum medium. This excretion did not take place in serum-free medium and was found to be stimulated by a nondialyzable, thermolabile component of human serum. Horse, chicken, calf, and rabbit serum also showed stimulation ability. The process of cholesterol excretion appears to be of general occurrence. It was found in both strains of cultured cells examined (mouse fibroblasts and lymphoblasts) and also in strips of rabbit aorta incubated in vitro.

scholarly journals “Humanized” Stem Cell Culture Techniques: The Animal Serum Controversy

2011 ◽  
Vol 2011 ◽  
pp. 1-14 ◽  
Author(s):  
Chandana Tekkatte ◽  
Gency Ponrose Gunasingh ◽  
K. M. Cherian ◽  
Kavitha Sankaranarayanan
Keyword(s):  
Cellular Therapy ◽  
Cultured Cells ◽  
Human Mesenchymal Stem Cells ◽  
The Body ◽  
Therapeutic Interventions ◽  
Post Transplantation ◽  
Cell Culture Techniques ◽  
Culture Techniques ◽  
Expansion Media

Cellular therapy is reaching a pinnacle with an understanding of the potential of human mesenchymal stem cells (hMSCs) to regenerate damaged tissue in the body. The limited numbers of these hMSCs in currently identified sources, like bone marrow, adipose tissue, and so forth, bring forth the need for theirin vitroculture/expansion. However, the extensive usage of supplements containing xenogeneic components in the expansion-media might pose a risk to the post-transplantation safety of patients. This warrants the necessity to identify and develop chemically defined or “humanized” supplements which would makein vitrocultured/processed cells relatively safer for transplantation in regenerative medicine. In this paper, we outline the various caveats associated with conventionally used supplements of xenogenic origin and also portray the possible alternatives/additives which could one day herald the dawn of a new era in the translation ofin vitrocultured cells to therapeutic interventions.

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