Baboon Peripheral Blood Mononuclear Cells Research Articles

Optimization of culture conditions for the derivation and propagation of baboon (Papio anubis) induced pluripotent stem cells.

Induced pluripotent stem cells (iPSCs) offer the possibility of cell replacement therapies using patient-matched cells to treat otherwise intractable diseases and debilitations. To successfully realize this potential, several factors must be optimized including i) selection of the appropriate cell type and numbers to transplant, ii) determination of the means of transplantation and the location into which the transplanted cells should be delivered, and iii) demonstration of the safety and efficacy of the cell replacement protocol to mitigate each targeted disease state. A majority of diseases or debilitations likely to be targeted by cell-based therapeutic approaches represent complex conditions or physiologies manifest predominantly in primates including humans. Nonhuman primates afford the most clinically relevant model system for biomedical studies and testing of cell-based therapies. Baboons have 92% genomic similarity with humans overall and especially significant similarities in their immunogenetic system, rendering this species a particularly valuable model for testing procedures involving cell transplants into living individuals. To maximize the utility of the baboon model, standardized protocols must be developed for the derivation of induced pluripotent stem cells from living adults and the long-term maintenance of these cells in culture. Here we tested four commercially available culture systems (ReproFF, mTeSR1, E8 and Pluristem) for competence to maintain baboon iPSCs in a pluripotent state over multiple passages, and to support the derivation of new lines of baboon iPSCs. Of these four media only Pluristem was able to maintain baboon pluripotency as assessed by morphological characteristics, immunocytochemistry and RT-qPCR. Pluristem also facilitated the derivation of new lines of iPSCs from adult baboon somatic cells, which had previously not been accomplished. We derived multiple iPS cell lines from adult baboon peripheral blood mononuclear cells cultured in Pluristem. These were validated by expression of the pluripotency markers OCT4, NANOG, SOX2, SSEA4 and TRA181, as well as the ability to differentiate into tissues from all three germ layers when injected into immunocompromised mice. These findings further advance the utility of the baboon as an ideal preclinical model system for optimizing iPS cell-based, patient-specific replacement therapies in humans.

Induction of porcine-specific regulatory T cells with high specificity and expression of IL-10 and TGF-β1 using baboon-derived tolerogenic dendritic cells.

Regulatory T cells (Treg) play an important role in maintenance of homeostasis in vivo. Treg application to alleviate allo-organ rejection is being studied extensively. However, natural Treg (nTreg) expansion in vitro is laborious and expensive. Antigen-specific Treg are more effective and require lower cell numbers than use of nTreg for immune control. The baboon, as a non-human primate experimental animal model, is widely used in xenotransplantation research. An effective method to generate baboon xeno-specific Treg would benefit research on immune tolerance in xenotransplantation using this model system. Baboon tolerogenic dendritic cells (tolDC) were generated in 3days from monocytes isolated from baboon peripheral blood mononuclear cells in medium supplemented with anti-inflammatory cytokines. After loading with porcine-specific (PS) in vitro-transcribed RNA (ivtRNA), tolDC were used to induce CD4+ T cells to become porcine-specific Treg (PSTreg) in cocultures supplemented with IL-2 and rapamycin for 10days. Anti-inflammatory and inflammatory cytokine expression was evaluated at the mRNA and protein levels in both baboon tolDC and PSTreg. Functional assays, suppression of activation markers on porcine-specific effector T cells (PSTeff) and inhibition of PSTeff proliferation, were used to test PSTreg specificity. TolDC generated with this method exhibited a tolerogenic phenotype, expressed CCR7 and produced high levels of IL-10 and TGF-β1, whereas IL-12p40 and IFN-γ were not expressed. PSTreg were successfully generated in cocultures of CD4+ T cells and PS ivtRNA-loaded tolDC. They exhibited a CD3+ CD4+ CD25+ CD127low/- CD45RAlow Foxp3+ phenotype and were characterized by high expression of IL-10 and TGF-β1 mRNA and protein. They showed upregulated expression of EBI3 and GARP mRNA. PSTreg exhibited highly suppressive effects toward PSTeff, secreting high amounts of IL-10 and TGF-β1 cytokine upon interaction with PSTeff and suppressing IFN-γ expression on PSTeff. In this study, a fast 3-day method to generate baboon-derived tolDC is provided that allows subsequent induction of PSTreg displaying high porcine-antigen specificity and expression of IL-10 and TGF-β1. Porcine-specific baboon Treg can be used in porcine solid organ or cell xenotransplantation studies through adoptive cell transfer into host baboons.

Ex vivo-expanded baboon CD39+ regulatory T cells prevent rejection of porcine islet xenografts in NOD-SCID IL-2rγ-/- mice reconstituted with baboon peripheral blood mononuclear cells.

A high immunosuppressive burden is required for long-term islet xenograft survival in non-human primates even using genetically modified donor pigs. We aimed to investigate the capacity of baboon regulatory T cells (Treg) to suppress islet xenograft rejection, thereby developing a potential immunoregulatory or tolerance therapy that could be evaluated in NHP models of xenotransplantation. Baboon Treg expanded with stimulation by porcine peripheral blood mononuclear cells (PBMC) were characterized by cell phenotyping and suppressive activity assays in vitro. Their function in vivo was evaluated in neonatal porcine islet cell clusters (NICC) transplanted NOD-SCID IL-2rγ-/- (NSG) mice receiving baboon PBMC alone or with expanded autologous Treg. The majority of expanded Treg coexpressed Foxp3 and CD39 and were highly suppressive of the baboon anti-pig xenogeneic T cell response in vitro. Reconstitution of mice with baboon PBMC alone resulted in NICC xenograft rejection within 35days. Cotransfer with baboon PBMC and Treg prolonged islet xenograft survival beyond 100days, correlating with Treg engraftment, intragraft CD39 and Foxp3 gene expression, and reduced graft infiltrating effector T cells and reduced interferon-γ production. Our data supports the capacity of ex vivo expanded CD39+ baboon Treg to suppress islet xenograft rejection in primatized mice, suggesting it haspotential as an adjunctive immunotherapy in preclinical NHP models of xenotransplantation.

The effect of Gal expression on pig cells on the human T‐cell xenoresponse

Lack of Gal expression on pig cells is associated with a reduced primate humoral immune response as well as a reduction in cytokine production by human cells in vitro. We investigated whether lack of Gal expression is associated with reduced human T-cell response in vitro. Peripheral blood mononuclear cells (PBMCs) were obtained from healthy humans and naïve baboons. Human CD4+ and CD8+ T cells were isolated. Porcine aortic endothelial cells (pAECs) were isolated from wild-type (WT) and α1,3-galactosyltransferase gene-knockout (GTKO) pigs. WT pAECs were treated with α-galactosidase, reducing Gal expression. Swine leukocyte antigen (SLA) class I and II expression on pAECs was measured, as was T-cell proliferation and cytokine production in response to pAECs. Reduced Gal expression on WT pAECs after α-galactosidase treatment was associated with reduced human PBMC proliferation (P<0.005). SLA class I and II expression on WT and GTKO pAECs was comparable. Human CD4+ and CD8+ T-cell proliferation was less against GTKO pAECs before (P<0.001) and after (P<0.01 and P<0.05, respectively) activation. Human and baboon PBMC proliferation was less against GTKO pAECs before (P<0.05) and after (P<0.01 and P<0.05, respectively) activation. Human PBMCs produced a comparable cytokine/chemokine response to WT and GTKO pAECs. However, there was less production of IFN-γ/TNF-α by CD4+ and IFN-γ/granzyme B/IP-10 by CD8+ T cells in response to GTKO pAECs. The absence of Gal on pig cells is associated with reduced human T-cell proliferation (and possibly selected cytokine production). Adaptive primate T-cell responses are likely to be reduced in GTKO xenograft recipients.

Recipient Tissue Factor Expression Is Associated With Consumptive Coagulopathy in Pig‐to‐Primate Kidney Xenotransplantation

Consumptive coagulopathy (CC) remains a challenge in pig-to-primate organ xenotransplantation (Tx). This study investigated the role of tissue factor (TF) expression on circulating platelets and peripheral blood mononuclear cells (PBMCs). Baboons (n = 9) received a kidney graft from pigs that were either wild-type (n = 2), alpha1,3-galactosyltransferase gene-knockout (GT-KO; n = 1) or GT-KO and transgenic for the complement-regulatory protein, CD46 (GT-KO/CD46, n = 6). In the baboon where the graft developed hyperacute rejection (n = 1), the platelets and PBMCs expressed TF within 4 h of Tx. In the remaining baboons, TF was detected on platelets on post-Tx day 1. Subsequently, platelet-leukocyte aggregation developed with formation of thrombin. In the six baboons with CC, TF was not detected on baboon PBMCs until CC was beginning to develop. Graft histopathology showed fibrin deposition and platelet aggregation (n = 6), but with only minor or no features indicating a humoral immune response (n = 3), and no macrophage, B or T cell infiltration (n = 6). Activation of platelets to express TF was associated with the initiation of CC, whereas TF expression on PBMCs was concomitant with the onset of CC, often in the relative absence of features of acute humoral xenograft rejection. Prevention of recipient platelet activation may be crucial for successful pig-to-primate kidney Tx.

Molecular pathways mediating differential responses to lipopolysaccharide between human and baboon arterial endothelial cells

1. Vascular inflammation plays a critical role in atherogenesis. Previously, we showed that baboon arterial endothelial cells (BAEC) were hyporesponsive to lipopolysaccharide (LPS) compared with human arterial endothelial cells (HAEC). 2. In the present study, we investigated mechanisms underlying differential responses between HAEC and BAEC to tumour necrosis factor (TNF)-alpha and LPS. 3. Both HAEC and BAEC responded similarly to TNF-alpha. However, BAEC showed retarded responses to LPS in expression of E-selectin, intercellular adhesion molecule-1, monocyte chemotactic protein-1 and interleukin-8 (P < 0.05). These changes were confirmed at the mRNA level. Tumour necrosis factor-alpha activated nuclear factor-kappaB members such as p50, p52, p65, c-rel and RelB in both HAEC and BAEC. In contrast, LPS activated p50 and p65 only in HAEC. Using microarray assays, we found that TNF receptor-associated factor 2 (TRAF-2), TNF receptor superfamily, member 1A-associated via death domain (TRADD) and nuclear factors such as nuclear factor of kappa in B-cells inhibitor, alpha (NFKBIA) and nuclear factor of kappa in B-cells inhibitor, beta (NFKBIB) were upregulated by LPS only in HAEC. Although the baseline expression of Toll-like receptor (TLR) 4 was low in both HAEC and BAEC, TNF-alpha activated TLR4 expression in both cell types. Although LPS increased TLR4 expression only in HAEC, human and baboon peripheral blood mononuclear cells exhibited similar TLR4 expression and response to LPS. Transfecting BAEC with TLR4/myeloid differentiation protein-2 overexpression vector conferred BAEC responsiveness to LPS. 4. The findings of the present study indicate that an altered TLR4 system may be responsible for the resistance of baboon endothelial cells to LPS. Given the importance of TLR4 in human immune responses and vascular diseases, the natural resistance of baboons to LPS/TLR4-initiated inflammation could make the baboon a valuable animal model in which to study how inflammation affects atherogenesis.

Atorvastatin Down-Regulates the Primate Cellular Response to Porcine Aortic Endothelial Cells In Vitro

Using mixed lymphocyte reaction (MLR), the effect of atorvastatin on proliferation of human and baboon peripheral blood mononuclear cells (PBMCs) and human CD4+ T cells in response to wild-type (WT) and alpha-1,3-galactosyltransferase gene-knockout (GTKO) porcine aortic endothelial cells (pAECs) was investigated. swine leukocyte antigen class-II (SLA II) expression on pAEC before and after porcine interferon gamma (pIFN-gamma) stimulation, and the effect of atorvastatin on this expression was assessed. Added to the MLR, atorvastatin reduced (i) the human PBMC response to unstimulated (P<0.05) and (ii) the human and baboon PBMC responses to stimulated (P<0.05) WT and GTKO pAEC. Atorvastatin treatment of human PBMC before MLR reduced their response to stimulated WT (P<0.05) and GTKO (P<0.05) pAEC. Stimulation of pAEC with pIFN-gamma increased SLA II expression 20- to 60-fold, which was down-regulated by atorvastatin. Atorvastatin treatment of stimulated pAEC before MLR reduced proliferation of human PBMC (P<0.05) and CD4+ T cells (P<0.05). Atorvastatin down-regulates the primate cellular xenoresponse, possibly through its antiproliferative effect on PBMCs and the reduction of SLA II on pAECs.

Initial experience with the human anti-human CD154 monoclonal antibody, ABI793, in pig-to-baboon xenotransplantation.

ABI793 (ABI) is a human monoclonal antibody (mAb) specific for human CD154. To assess the suitability of ABI for baboon transplantation studies, we carried out in vitro studies to determine ABI's reactivity with baboon cells expressing CD154, performed in vivo pharmacokinetic studies in two baboons, and tested the effect of ABI administration on elicited antibody production in two baboons undergoing either pig hematopoietic progenitor cell (PBPC) or heterotopic heart transplantation. In vitro: Baboon peripheral blood mononuclear cells were activated in vitro to upregulate CD154, and binding of ABI to CD154 was measured by flow cytometry. In vivo: Serum levels of ABI were measured immediately before and 15 min after the intravenous administration of ABI (20 mg/kg) to two baboons over 28 days. Subsequently, ABI (25 mg/kg on days 0, 1, 4 and 7, and then 20 mg/kg every 5 days) was included in the immunosuppressive regimen in two pig-to-baboon transplants (PBPC or heart transplantation). In vitro: ABI was almost non-reactive to baboon T cells before stimulation, but bound to activated T cells. In vivo: In the pharmacokinetic study, trough levels of ABI (before the next dose) ranged between 190 and 580 microg/ml, and the estimated half-life was 10-15 days. There was no apparent toxicity. Following pig PBPC or heart transplantation, no elicited antibody was detected while ABI was being administered or during several weeks of follow-up. ABI functions in baboons, is well-tolerated, and prevents an elicited antibody response to pig antigens.

Absence of humoral and cellular alloreactivity in baboons sensitized to pig antigens.

to study whether sensitization to pig antigens results in humoral and/or cellular sensitization to alloantigens in baboons, and thus increases the risks of organ allotransplantation after xenotransplantation. Serum from baboons that were naive (n = 4), sensitized to Gal alpha 1,3Gal (Gal) antigens (n = 2), or sensitized to Gal + non-Gal pig antigens (n = 2) were tested by flow cytometry for the presence of immunoglobulin G (IgG) and IgM antibodies that bind to pig or baboon peripheral blood mononuclear cells (PBMC). Two allosensitized baboons were used as positive controls. The same 10 sera were tested in a complement-mediated cytotoxicity assay to detect cytotoxic antibodies against pig, allo and self-PBMC. The T-cell responses of the same baboons to allogeneic and pig PBMC stimulators in mixed lymphocyte reaction (MLR) were studied. All baboon sera contained cytotoxic antibodies that bound to pig PBMC. Binding and cytotoxicity were higher in xenosensitized baboons, particularly in those sensitized to Gal + non-Gal antigens (P < 0.001). None of the naive or xenosensitized baboon sera bound to baboon PBMC. Serum from allosensitized baboons showed anti-baboon IgG and IgM binding, but there was no increase in binding to pig PBMC or in cytotoxicity to pig cells. The MLR response to pig stimulators in baboons sensitized to non-Gal pig antigens was greater than that of naive or Gal-sensitized baboons (P < 0.001), but there was no increase in the response to baboon cells. In baboons, no in vitro evidence that a previous pig xenograft might endanger the outcome of a subsequent allograft was documented.

Increased virus replication and virulence after serial passage of human immunodeficiency virus type 2 in baboons.

Similar to human immunodeficiency virus type 1 (HIV-1) infection of humans, the natural history of HIV-2 infection in baboons (Papio cynocephalus) is a slow and chronic disease that generally takes several years before an AIDS-like condition develops. To shorten the amount of time to the development of disease, we performed five serial passages of HIV-2(UC2) in baboons by using blood and bone marrow samples during the acute phase of infection when viral loads were at high levels. After these serial passages, virus levels in plasma, peripheral blood mononuclear cells (PBMC) and lymphatic tissues in the acutely infected baboons were increased. Within 1 year of the HIV-2 infection, all of the inoculated baboons showed specific signs of AIDS-related disease progression within the lymphatic tissues, such as vascular proliferation and lymphoid depletion. The HIV-2(UC2) recovered after four serial passages showed increased kinetics of viral replication in baboon PBMC and cytopathicity. This study suggests that the HIV-2 isolate recovered after several serial passages in baboons will be useful in future studies of AIDS pathogenesis and vaccine development by using this animal model.

EFFECTS ON HUMAN AND NONHUMAN PRIMATE IMMUNE RESPONSE OF A NEW RAT ANTI-CD2 MONOCLONAL ANTIBODY1

Nonhuman primate models are highly clinically relevant in transplantation. The development of immunosuppressive tools or a tolerogenic regimen for primate models therefore represents an important goal of transplantation immunological research. Hence, we have developed a rat monoclonal antibody (mAb) that recognizes the CD2 molecule (LO-CD2b) on both human and nonhuman primate cells. The LO-CD2b mAb has been characterized by flow cytometry, E-rosetting inhibition, and Western blotting. In vitro inhibition of immune responses by LO-CD2b was assessed after both mitogenic and allogeneic stimulation in mixed lymphocyte reactions (MLR). Several LO-CD2b dose and time responses were tested. In vivo, peripheral and lymph node T-cell depletion was examined both by flow cytometry and immunohistology in 10 baboons that received intravenous injection of LO-CD2b at different doses and time courses. Xenosensitization (anti-rat) was assessed by ELISA. Renal allograft survival was followed in two baboons treated with iterative LO-CD2b injections. In vitro, LO-CD2b binds a lymphocyte antigenic determinant of 52 kDa that is recognized by other well-characterized anti-CD2 mAbs (T11, Leu5b). LO-CD2b recognized natural killer CD2+ cells. Administration of 200 ng/ml LO-CD2b almost completely inhibited human and baboon mitogenic stimulation. Allogeneic baboon and human MLR were completely inhibited by the addition of LO-CD2b (at 312 ng/ml) on the day of the initiation of culture; when added after 1 or 2 days, LO-CD2b still provided a significant MLR inhibition (>50%). Incubation of LO-CD2b with baboon peripheral blood mononuclear cells produced very low cytokine levels (interferon-y, tumor necrosis factor-alpha, interleukin 2). In secondary MLR, baboon peripheral blood mononuclear cells previously incubated with LO-CD2b were unable to respond to a second allogeneic stimulation but were able to react to mitogens. In vivo, within the first hour after LO-CD2b injection (at 0.15, 0.5, and 2 mg/kg), an 85-90% peripheral depletion of CD2+ cells was observed. A partial T-cell depletion in inguinal lymph nodes was seen after 1 week. The mechanism of peripheral T-cell depletion could have been antibody-dependent cell cytotoxicity or opsonization but was complement independent. Iterative LO-CD2b injections (12 days at 0.35 mg/kg) slightly prolonged the renal allograft survival in two baboons. LO-CD2b is a nonactivating rat anti-CD2 mAb able to strongly inhibit both mitogenic and allogeneic responses in human and nonhuman primates. In vivo, LO-CD2b provides a rapid peripheral T-cell depletion, which is reversible within days after the cessation of injections. This rat mAb represents a very important tool for in vivo experimental investigation in nonhuman primates because it similarly reacts against human T cells in vitro.

Infection of baboons with a simian immunodeficiency virus/HIV-1 chimeric virus constructed with an HIV-1 Thai subtype E envelope.

To construct an infectious chimeric simian immunodeficiency virus/HIV-1 (SHIV) with the envelope of a Thai subtype E HIV-1 strain for use in a non-human primate model. A novel SHIV genome was derived using the sequences of the ectodomain of the envelope gene from the Thai subtype E strain, HIV-1(9466). This SHIV (SHIV9466.33) was recovered by cocultivation from human peripheral blood mononuclear cells (PBMC) after transfection of human rhabdosarcoma cells. Rhesus macaque and baboon PBMC were screened in vitro for susceptibility to infection with SHIV9466.33. After successful infection of baboon PBMC, four animals were inoculated intravenously with SHIV9466.33 and monitored for plasma viral RNA, virus isolation from the PBMC, seroconversion, T-cell subsets, and signs of disease. SHIV9466.33 was able to infect PBMC from 12 out of 14 baboons. All four of the baboons selected for in vivo inoculation became infected. Peak plasma viral RNA levels of 8000 to 700,000 RNA copies/ml were measured at 2 weeks post-inoculation. Virus was isolated from the PBMC of all four baboons during acute infection, and all seroconverted. Although transient declines in CD4+ T-cells were observed during early infection, CD4+ levels remained within normal ranges thereafter. In contrast, in vitro cultures of PBMC of four rhesus macaques were not susceptible to infection with SHIV9466.33. SHIV9466.33 containing an HIV-1 subtype E envelope displayed tropism for baboon PBMC but not for rhesus macaque PBMC. This chimeric virus established infection and induced antiviral antibodies in baboons inoculated by the intravenous route with cell-free virus. Thus, infection of baboons with SHIV9466.33 will serve as an important animal model for future studies of HIV-1 vaccine efficacy.

Transient virus infection and pathogenesis of a new HIV type 2 isolate, UC12, in baboons.

We have previously shown that baboons (Papio cynocephalus) can be persistently infected with HIV-2 and some baboons progress to an AIDS-like disease with a CD4+ T cell decline, cachexia, alopecia, and Kaposi's sarcoma-like fibromatosis. In this study, we found that a new virus isolate, HIV-2UC12, replicated to high levels in baboon peripheral blood mononuclear cells (PBMCs) in vitro. Three baboons were subsequently inoculated and had plasma viral RNA loads that peaked between 15,000 and 7000 copies/ml at 2 weeks postinfection. Virus was isolated from the PBMCs for up to 6 months. Although PBMCs were subsequently virus culture negative, virus could be recovered from the spleen, lymph nodes, and tonsils, indicating that HIV-2 was sequestered within these lymphoid tissues. HIV-2-associated pathology included follicular lysis, vascular proliferation, and lymphoid depletion. This study indicated that HIV-2UC12 infection in baboons can cause HIV-associated pathological abnormalities within the lymphatic tissues and that the high level of HIV-2UC12 replication in vitro was not predictive of replication in vivo.

CD8+ cells from HIV-2-infected baboons control HIV replication.

To analyze the CD8+ cell antiviral immune response in HIV-2-infected baboons. Baboons were infected with clinical isolates of HIV-2, CD8+ cells were isolated from phytohemagglutinin (PHA)-stimulated baboon peripheral blood mononuclear cells (PBMC). These cells were cultured with PHA-stimulated CD4+ cells acutely infected with HIV-2 at several CD8+:CD4+ cell ratios. Control of HIV-2 replication was determined by comparing peak levels of HIV-2 replication in fluids from CD8+:CD4+ cell cocultures with those in fluids from infected CD4+ cells cultured alone. CD8+ cells from HIV-2-infected baboons inhibited HIV-2 replication in acutely infected autologous CD4+ cells to a significantly greater extent than did CD8+ cells from uninfected baboons (P = 0.0001). At the beginning of the acute phase of HIV-2 infection, CD8+ cells showed either a transient reduction or loss in the antiviral activity. In some cases the CD8+ cell response enhanced HIV-2 replication. Subsequently, the strength of the CD8+ cell antiviral activity increased concomitant with a decrease in the HIV-2 load in the PBMC. Suppression of HIV replication could be demonstrated with filtered fluid from CD8+ cells. Other studies indicated that infected CD4+ cells are lost during coculture of CD8+ cells with infected CD4+ cells. CD8+ cells of HIV-2-infected baboons develop substantial anti-HIV-2 activity following HIV-2 infection, which may account in part for the low frequency of pathogenesis in HIV-2-infected baboons. Studies to elucidate the mechanism of this CD8+ cell antiviral activity suggest that it is mediated in part by a soluble antiviral factor, but primarily in association with the loss of infected CD4+ cells.

Characterization of Infectious Type D Retrovirus from Baboons

Infectious virus resembling type D simian retrovirus (SRV) was isolated from Ethiopian baboons ( Papio cynocephalus) (SRV-Pc) housed at the University of Washington Regional Primate Research Center. When baboon peripheral blood mononuclear cells (PBMC) or tissues were cocultured with the H-9 human T-cell line or the Raji human B-cell line, large multinucleated syncytia positive for SRV-2 antigens were observed microscopically. Immunoblot analysis of purified SRV-Pc from cell culture supernatants demonstrated that the viral core and envelope proteins reacted with rabbit anti-SRV-2 serum. Fresh PBMC and cocultured cells were positive by polymerase chain reaction using two different sets of SRV-2 primers. Preliminary sequence analysis of two separate isolates from portions of the SRV-Pc p27 and gp20 regions revealed homology with SRV-1, SRV-2, and Mason-Pfizer monkey virus. The homologies in the p27 segment were 91-94% and the homologies in the gp20 segment were 72-75%.

Mitogenic responses of baboon ( [formula omitted]) peripheral blood mononuclear cells. A possible role for monocytes.

Removal of carbonyl iron-adherent/phagocytic cells from baboon peripheral blood mononuclear cells generally resulted in a depressed blastogenic response to both pokeweed mitogen and concanavalin A. In the majority of cases, no alteration in dose requirement nor shift in kinetics was apparent. Staining for peroxidase activity indicated a reduced proportion of monocytes in the population of cells treated with iron. Therefore, the results of this study strongly suggest a potentiating role for monocytes in lectin stimulation of baboon lymphocytes.

Micro-system for the primary immunization of baboon ( papio cynocephalus) peripheral blood mononuclear cells with sheep erythrocytes in vitro

A micro-culture system for the stimulation of baboon peripheral blood mononuclear cells (PBMC) with sheep erythrocytes (SRBC) in vitro was established. PBMC cultures in 96-well microtiter plates were maintained in 300 μl of culture medium containing a murine mixed lymphocyte reaction (MLR) supernatant and SRBC. Cultures of 7.5 × 10 5 PBMC and 6 × 10 6 SRBC resulted in the highest anti-SRBC plaque-forming cell (PFC) response. It was also determined that the presence of 50 μl of murine MLR supernatant was required for optimal PFC generation and that the cultures required supplementation with nutrient cocktail 3 days post-immunization. Although PFC were detectable on days 4–8, the maximum expression of PFC occurred on day 7.