Loss of T cell responses following long-term cryopreservation.

Author(s): Owen RE, Sinclair E, Emu B, Heitman JW, Hirschkorn DF, Epling CL, Tan QX, Custer B, Harris JM, Jacobson MA, McCune JM, Martin JN, Hecht FM, Deeks SG, Norris PJ

Publication: J Immunol Methods, 2007, Vol. 326, Page 93-115

PubMed ID: 17707394 PubMed Review Paper? No

Purpose of Paper

Conclusion of Paper

Studies

1. Study Purpose

   The purpose of this study was to determine the effects of cryopreservation, B-LCL addition and cell resting on CD4+ and CD8+ response to stimulation in PBMC from HIV positive and negative individuals. Cells were stored at -135°C.

   Summary of Findings:

   After long-term cryopreservation, 94% of PBMC were viable. Long-term cryopreservation caused decreased CD4+ T cell response to p55 protein (0.29%; p=0.023), p55 GAG peptide (p=0.043) and CMV lysate (0.59%; p=0.038) and a non-significant trend toward decreased response to HIV peptides (0.87%; p=0.105) and increased response to MN (0.069%; p=0.806) in HIV positive specimens. Cryopreservation also caused decreased CD8+ T cell responses to p55 protein (1.348%; P=0.004) and CMV lysate (1.637%; p=0.041) and a non-significant decline in response to HIV peptide (0.281%; p=0.187) and MN (0.254% p=0.187) in HIV positive specimens. The loss of p55 GAG peptide response of CD4+ T cells in HIV positive specimens increased with longer storage. A prepulse of B-LCL did not significantly restore CD4+ or CD8+ response to p55 protein stimulation in the 4 individuals tested, but some changes in individual specimens were noted. Allowing the specimens to rest overnight following thaw also did not restore CD4+ or CD8+ response in HIV positive individuals. In HIV negative individuals, CD4+ and CD8+ responses to CMV pp65 were not substantially altered and showed less variability after storage.

   Biospecimens

   • Cell - Blood

   Preservative Types

   • Frozen

   Diagnoses:

   • AIDS/HIV-related
   • Normal

   Platform:

   Analyte	Technology Platform
   Cell count/volume	Flow cytometry

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Storage	Storage duration	0 days 0-170 days More than 300 days
   Preaquisition	Diagnosis/ patient condition	HIV positive HIV negative
   Biospecimen Aliquots and Components	Blood and blood products	Peripheral blood mononuclear cells

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2. Study Purpose

   The purpose of this study was to determine if cryopreservation (337-591 d) effects T cell phenotype and apoptosis in HIV positive PBMC specimens. Cells were stored at -135°C.

   Summary of Findings:

   Cryopreservation caused a decrease in naive CD4+ (p=0.0003) and CD8+ T cells (p=0.044) and a increase in CD4+ effector T cells (p=0.03) and CD8+ central memory T cells (p=0.041), but no change in CD4+ central memory T cells or CD8+ effector T cells was observed. However, variability within individuals in CD8+ effector T cells was very high. Further after long term cryopreservation CD4+ T cells stimulated with staphylococcal enterotoxin B or p55 protein had higher levels of caspase 3 staining indicating increased apoptosis. No effect of cryopreservation on apoptosis was observed in CD8+ T cells, or in unstimulated or CMV lysate stimulated CD4+ T cells.

   Biospecimens

   • Cell - Blood

   Preservative Types

   • Frozen

   Diagnoses:

   • AIDS/HIV-related

   Platform:

   Analyte	Technology Platform
   Cell count/volume	Flow cytometry

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Biospecimen Aliquots and Components	Blood and blood products	Peripheral blood mononuclear cells
   Storage	Storage duration	0 days 331-537 days

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