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1.
J Infect Dis ; 226(3): 521-527, 2022 08 26.
Artículo en Inglés | MEDLINE | ID: mdl-35290467

RESUMEN

Plasmodium falciparum-infected erythrocytes that display the variant surface antigen VAR2CSA bind chondroitin sulfate A (CSA) to sequester in placental intervillous spaces, causing severe sequelae for mother and offspring. Here, we establish a placental malaria (PM) monkey model. Pregnant Aotus infected with CSA-binding P. falciparum CS2 parasites during the third trimester developed pronounced sequestration of late-stage parasites in placental intervillous spaces that express VAR2CSA and bind specifically to CSA. Similar to immune multigravid women, a monkey infected with P. falciparum CS2 parasites over successive pregnancies acquired antibodies against VAR2CSA, with potent functional activity that was boosted upon subsequent pregnancy infections. Aotus also developed functional antibodies after multiple acute PM episodes and subsequent VAR2CSA immunization. In summary, P. falciparum infections in pregnant Aotus monkeys recapitulate all the prominent features of human PM infection and immunity, and this model can be useful for basic mechanistic studies and preclinical studies to qualify candidate PM vaccines. Clinical Trials Registration: NCT02471378.


Asunto(s)
Malaria Falciparum , Malaria , Complicaciones Parasitarias del Embarazo , Animales , Anticuerpos Antiprotozoarios , Antígenos de Protozoos , Aotidae , Sulfatos de Condroitina , Eritrocitos , Femenino , Humanos , Placenta , Plasmodium falciparum , Embarazo
2.
Proc Natl Acad Sci U S A ; 115(49): 12513-12518, 2018 12 04.
Artículo en Inglés | MEDLINE | ID: mdl-30455312

RESUMEN

Concerns about malaria parasite resistance to treatment with artemisinin drugs (ARTs) have grown with findings of prolonged parasite clearance t1/2s (>5 h) and their association with mutations in Plasmodium falciparum Kelch-propeller protein K13. Here, we describe a P. falciparum laboratory cross of K13 C580Y mutant with C580 wild-type parasites to investigate ART response phenotypes in vitro and in vivo. After genotyping >400 isolated progeny, we evaluated 20 recombinants in vitro: IC50 measurements of dihydroartemisinin were at similar low nanomolar levels for C580Y- and C580-type progeny (mean ratio, 1.00; 95% CI, 0.62-1.61), whereas, in a ring-stage survival assay, the C580Y-type progeny had 19.6-fold (95% CI, 9.76-39.2) higher average counts. In splenectomized Aotus monkeys treated with three daily doses of i.v. artesunate, t1/2 calculations by three different methods yielded mean differences of 0.01 h (95% CI, -3.66 to 3.67), 0.80 h (95% CI, -0.92 to 2.53), and 2.07 h (95% CI, 0.77-3.36) between C580Y and C580 infections. Incidences of recrudescence were 57% in C580Y (4 of 7) versus 70% in C580 (7 of 10) infections (-13% difference; 95% CI, -58% to 35%). Allelic substitution of C580 in a C580Y-containing progeny clone (76H10) yielded a transformant (76H10C580Rev) that, in an infected monkey, recrudesced regularly 13 times over 500 d. Frequent recrudescences of ART-treated P. falciparum infections occur with or without K13 mutations and emphasize the need for improved partner drugs to effectively eliminate the parasites that persist through the ART component of combination therapy.


Asunto(s)
Antimaláricos/farmacología , Artemisininas/farmacología , Malaria Falciparum/parasitología , Plasmodium falciparum/efectos de los fármacos , Animales , Aotidae , Cruzamientos Genéticos , Resistencia a Medicamentos , Regulación de la Expresión Génica , Mutación , Proteínas Protozoarias/genética , Proteínas Protozoarias/metabolismo
3.
Xenotransplantation ; 26(2): e12465, 2019 03.
Artículo en Inglés | MEDLINE | ID: mdl-30290025

RESUMEN

A combination of genetic manipulations of donor organs and target-specific immunosuppression is instrumental in achieving long-term cardiac xenograft survival. Recently, results from our preclinical pig-to-baboon heterotopic cardiac xenotransplantation model suggest that a three-pronged approach is successful in extending xenograft survival: (a) α-1,3-galactosyl transferase (Gal) gene knockout in donor pigs (GTKO) to prevent Gal-specific antibody-mediated rejection; (b) transgenic expression of human complement regulatory proteins (hCRP; hCD46) and human thromboregulatory protein thrombomodulin (hTBM) to avoid complement activation and coagulation dysregulation; and (c) effective induction and maintenance of immunomodulation, particularly through co-stimulation blockade of CD40-CD40L pathways with anti-CD40 (2C10R4) monoclonal antibody (mAb). Using this combination of manipulations, we reported significant improvement in cardiac xenograft survival. In this study, we are reporting the survival of cardiac xenotransplantation recipients (n = 3) receiving xenografts from pigs without the expression of hTBM (GTKO.CD46). We observed that all grafts underwent rejection at an early time point (median 70 days) despite utilization of our previously reported successful immunosuppression regimen and effective control of non-Gal antibody response. These results support our hypothesis that transgenic expression of human thrombomodulin in donor pigs confers an independent protective effect for xenograft survival in the setting of a co-stimulation blockade-based immunomodulatory regimen.


Asunto(s)
Rechazo de Injerto/inmunología , Supervivencia de Injerto/inmunología , Xenoinjertos/inmunología , Trombomodulina/inmunología , Trasplante Heterólogo , Animales , Animales Modificados Genéticamente , Técnicas de Inactivación de Genes , Rechazo de Injerto/genética , Supervivencia de Injerto/genética , Trasplante de Corazón/métodos , Terapia de Inmunosupresión/métodos , Inmunosupresores/farmacología , Porcinos , Trasplante Heterólogo/métodos
4.
Xenotransplantation ; 25(2): e12379, 2018 03.
Artículo en Inglés | MEDLINE | ID: mdl-29250828

RESUMEN

BACKGROUND: CD4+CD25Hi FoxP3+ T (Treg) cells are a small subset of CD4+ T cells that have been shown to exhibit immunoregulatory function. Although the absolute number of Treg cells in peripheral blood lymphocytes (PBL) is very small, they play an important role in suppressing immune reactivity. Several studies have demonstrated that the number of Treg cells, rather than their intrinsic suppressive capacity, may contribute to determining the long-term fate of transplanted grafts. In this study, we analyzed Treg cells in PBL of long-term baboon recipients who have received genetically modified cardiac xenografts from pig donors. METHODS: Heterotopic cardiac xenotransplantation was performed on baboons using hearts obtained from GTKO.hCD46 (n = 8) and GTKO.hCD46.TBM (n = 5) genetically modified pigs. Modified immunosuppression regimen included antithymocyte globulin (ATG), anti-CD20, mycophenolate mofetil (MMF), cobra venom factor (CVF), and costimulation blockade (anti-CD154/anti-CD40 monoclonal antibody). FACS analysis was performed on PBLs labeled with anti-human CD4, CD25, and FoxP3 monoclonal antibodies (mAb) to analyze the percentage of Treg cells in six baboons that survived longer than 2 months (range: 42-945 days) after receiving a pig cardiac xenograft. RESULTS: Total WBC count was low due to immunosuppression in baboons who received cardiac xenograft from GTKO.hCD46 and GTKO.hCD46.hTBM donor pigs. However, absolute numbers of CD4+CD25Hi FoxP3 Treg cells in PBLs of long-term xenograft cardiac xenograft surviving baboon recipients were found to be increased (15.13 ± 1.50 vs 7.38 ± 2.92; P < .018) as compared to naïve or pre-transplant baboons. Xenograft rejection in these animals was correlated with decreased numbers of regulatory T cells. CONCLUSION: Our results suggest that regulatory T (Treg) cells may contribute to preventing or delaying xenograft rejection by controlling the activation and expansion of donor-reactive T cells, thereby masking the antidonor immune response, leading to long-term survival of cardiac xenografts.


Asunto(s)
Trasplante de Corazón , Xenoinjertos/inmunología , Linfocitos T Reguladores/inmunología , Tiempo , Trasplante Heterólogo , Animales , Animales Modificados Genéticamente , Factores de Transcripción Forkhead/inmunología , Rechazo de Injerto/tratamiento farmacológico , Rechazo de Injerto/inmunología , Supervivencia de Injerto/inmunología , Trasplante de Corazón/métodos , Tolerancia Inmunológica , Terapia de Inmunosupresión/métodos , Inmunosupresores/farmacología , Papio , Porcinos , Trasplante Heterólogo/métodos
5.
Xenotransplantation ; 24(6)2017 11.
Artículo en Inglés | MEDLINE | ID: mdl-28940570

RESUMEN

BACKGROUND: Innovations in transgenic technology have facilitated improved xenograft survival. Additional gene expression appears to be necessary to overcome the remaining immune and biologic incompatibilities. We report for the first time the novel use of six-gene modifications within a pig-to-baboon cardiac xenotransplantation model. METHODS: Baboons (8-15 kg) underwent heterotopic cardiac transplantation using xenografts obtained from genetically engineered pigs. Along with previously described modifications (GTKO, hCD46), additional expression of human transgenes for thromboregulation (endothelial protein C receptor, tissue factor pathway inhibitor, thrombomodulin), complement inhibition (decay accelerating factor), and cellular immune suppression (hCD39, hCD47) was used. Immunosuppression consisted of targeted T-cell and B-cell depletion and conventional anti-rejection agents. RESULTS: Heterotopic cardiac transplantations were performed without complication. Flow cytometry and immunohistochemistry on donor biopsies confirmed transgenic phenotype. In contrast to the prior three-gene generation, significant coagulopathy or consumptive thrombocytopenia has not been observed in the six-gene cohort. As a result, these recipients have experienced decreased bleeding-related complications. Pro-inflammatory responses also appear to be mitigated based on cytokine analysis. Baboons survived the critical 30-day post-operative period when mortality has historically been highest, with no evidence of graft rejection. CONCLUSIONS: The inclusion of additional human genes in genetically engineered pigs appears to confer superior xenograft outcomes. Introduction of these genes has not been associated with adverse outcomes. This multifactorial approach to genetic engineering furthers the prospect of long-term cardiac xenograft survival and subsequent clinical application.


Asunto(s)
Rechazo de Injerto/inmunología , Trasplante de Corazón , Xenoinjertos/inmunología , Inmunosupresores/farmacología , Trasplante Heterólogo , Animales , Animales Modificados Genéticamente/genética , Animales Modificados Genéticamente/inmunología , Supervivencia de Injerto/inmunología , Trasplante de Corazón/métodos , Terapia de Inmunosupresión/métodos , Papio/metabolismo , Papio hamadryas , Porcinos , Trasplante Heterólogo/métodos , Trasplante Heterotópico/métodos
6.
J Med Primatol ; 45(6): 312-317, 2016 12.
Artículo en Inglés | MEDLINE | ID: mdl-27477611

RESUMEN

BACKGROUND: Klebsiella pneumoniae can be a serious pathogen in non-human primates, particularly Neotropical monkeys. METHODS: During a K. pneumoniae outbreak in an owl monkey research colony, 13 K. pneumoniae isolates were DNA fingerprinted by automated repetitive extragenic palindromic-polymerase chain reaction and the profiles compared to isolates obtained from other non-human primate species during the same time period and isolates from previous outbreaks. RESULTS: Eleven different types of K. pneumoniae were circulating in the owl monkey colony at the time of the outbreak. When comparing owl monkey isolates relatedness to previous colony outbreak isolates and squirrel monkey and capuchin monkey isolates, all were different. CONCLUSIONS: These results agree with recent reports where K. pneumoniae nosocomial isolates in hospital settings can have high genetic diversity, and multiple strains can be circulating simultaneously. This potential genetic diversity should be considered when designing strategies for controlling K. pneumoniae outbreaks in captive non-human primate colonies.


Asunto(s)
Aotidae , Brotes de Enfermedades , Variación Genética , Infecciones por Klebsiella/epidemiología , Klebsiella pneumoniae/genética , Enfermedades de los Monos/epidemiología , Animales , Animales de Laboratorio , Femenino , Infecciones por Klebsiella/microbiología , Masculino , Enfermedades de los Monos/microbiología
7.
Xenotransplantation ; 21(1): 35-45, 2014.
Artículo en Inglés | MEDLINE | ID: mdl-24164510

RESUMEN

BACKGROUND: Recently, we have shown that an immunosuppression regimen including costimulation blockade via anti-CD154 antibody significantly prolongs the cardiac xenograft survival in a GTKO.hCD46Tg pig-to-baboon heterotopic xenotransplantation model. Unfortunately, many coagulation disorders were observed with the use of anti-CD154 antibody, and recipient survival was markedly reduced by these complications. MATERIAL AND METHODS: In this experiment, we replaced anti-CD154 antibody with a more clinically acceptable anti-CD40 antibody while keeping the rest of the immunosuppressive regimen and the donor pig genetics the same. This was carried out to evaluate the antibody's role in xenograft survival and prevention of coagulopathies. Two available clones of anti-CD40 antibody were tested. One mouse anti-human CD40 antibody, (clone 3A8), activated B lymphocytes in vitro and only modestly suppressed antibody production in vivo. Whereas a recombinant mouse non-human primate chimeric raised against macaque CD40, (clone 2C10R4), blocked B-cell activation in vitro and completely blocked antibody production in vivo. RESULTS: The thrombotic complications seen with anti-CD154 antibody were effectively avoided but the graft survival, although extended, was not as prolonged as observed with anti-CD154 antibody treatment. The longest survival for the 3A8 antibody group was 27 days, and the longest graft survival in the 2C10R4 antibody group was 146 days. All of the grafts except two rejected and were explanted. Only two recipient baboons had to be euthanized due to unrelated complications, and the rest of the baboons remained healthy throughout the graft survival period or after graft explantation. In contrast to our anti-CD 154 antibody-treated baboons, the non-Gal antibody levels started to rise after B cells made their appearance around 8 weeks post-transplantation. CONCLUSIONS: Anti-CD40 antibody at the current dose does not induce any coagulopathies but while effective, had reduced efficacy to induce similar long-term graft survival as with anti-CD154 antibody perhaps due to ineffective control of B-cell function and antibody production at the present dose. More experiments are required to determine antibody affinity and effective dose for inducing long-term cardiac xenograft survival.


Asunto(s)
Anticuerpos/inmunología , Antígenos CD40/inmunología , Rechazo de Injerto/inmunología , Supervivencia de Injerto/inmunología , Animales , Animales Modificados Genéticamente , Formación de Anticuerpos , Linfocitos B/inmunología , Ligando de CD40/inmunología , Rechazo de Injerto/prevención & control , Xenoinjertos , Inmunosupresores/farmacología , Papio , Sus scrofa , Porcinos , Trasplante Heterólogo/métodos
8.
Lab Anim (NY) ; 52(12): 315-323, 2023 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-37932470

RESUMEN

Placental malaria vaccines (PMVs) are being developed to prevent severe sequelae of placental malaria (PM) in pregnant women and their offspring. The leading candidate vaccine antigen VAR2CSA mediates parasite binding to placental receptor chondroitin sulfate A (CSA). Despite promising results in small animal studies, recent human trials of the first two PMV candidates (PAMVAC and PRIMVAC) generated limited cross-reactivity and cross-inhibitory activity to heterologous parasites. Here we immunized Aotus nancymaae monkeys with three PMV candidates (PAMVAC, PRIMVAC and ID1-ID2a_M1010) adjuvanted with Alhydrogel, and exploited the model to investigate boosting of functional vaccine responses during PM episodes as well as with nanoparticle antigens. PMV candidates induced high levels of antigen-specific IgG with significant cross-reactivity across PMV antigens by enzyme-linked immunosorbent assay. Conversely, PMV antibodies recognized native VAR2CSA and blocked CSA adhesion of only homologous parasites and not of heterologous parasites. PM episodes did not significantly boost VAR2CSA antibody levels or serum functional activity; nanoparticle and monomer antigens alike boosted serum reactivity but not functional activities. Overall, PMV candidates induced functional antibodies with limited heterologous activity in Aotus monkeys, similar to responses reported in humans. The Aotus model appears suitable for preclinical downselection of PMV candidates and assessment of antibody boosting by PM episodes.


Asunto(s)
Vacunas contra la Malaria , Malaria Falciparum , Malaria , Animales , Humanos , Femenino , Embarazo , Placenta/parasitología , Malaria Falciparum/prevención & control , Malaria Falciparum/parasitología , Plasmodium falciparum , Antígenos de Protozoos , Anticuerpos Antiprotozoarios , Malaria/prevención & control , Aotidae , Inmunidad
9.
Front Immunol ; 13: 994498, 2022.
Artículo en Inglés | MEDLINE | ID: mdl-36605190

RESUMEN

Tissue resident (TR) immune cells play important roles in facilitating tissue homeostasis, coordinating immune responses against infections and tumors, and maintaining immunological memory. While studies have shown these cells are distinct phenotypically and functionally from cells found in the peripheral blood (PB), the clonal relationship between these populations across tissues has not been comprehensively studied in primates or humans. We utilized autologous transplantation of rhesus macaque hematopoietic stem and progenitor cells containing high diversity barcodes to track the clonal distribution of T, B, myeloid and natural killer (NK) cell populations across tissues, including liver, spleen, lung, and gastrointestinal (GI) tract, in comparison with PB longitudinally post-transplantation, in particular we focused on NK cells which do not contain endogenous clonal markers and have not been previously studied in this context. T cells demonstrated tissue-specific clonal expansions as expected, both overlapping and distinct from blood T cells. In contrast, B and myeloid cells showed a much more homogeneous clonal pattern across various tissues and the blood. The clonal distribution of TR NK was more heterogenous between individual animals. In some animals, as we have previously reported, we observed large PB clonal expansions in mature CD56-CD16+ NK cells. Notably, we found a separate set of highly expanded PB clones in CD16-CD56- (DN) NK subset that were also contributing to TR NK cells in all tissues examined, both in TR CD56-CD16+ and DN populations but absent in CD56+16- TR NK across all tissues analyzed. Additionally, we observed sets of TR NK clones specific to individual tissues such as lung or GI tract and sets of TR NK clones shared across liver and spleen, distinct from other tissues. Combined with prior functional data that suggests NK memory is restricted to liver or other TR NK cells, these clonally expanded TR NK cells may be of interest for future investigation into NK cell tissue immunological memory, with implications for development of NK based immunotherapies and an understanding of NK memory.


Asunto(s)
Células Asesinas Naturales , Células Mieloides , Animales , Células Clonales , Macaca mulatta
10.
Neurooncol Adv ; 2(1): vdaa005, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32309806

RESUMEN

BACKGROUND: Epigenetic modifiers are being investigated for a number of CNS malignancies as tumor-associated mutations such as isocitrate dehydrogenase mutations (IDH1/IDH2) and H3K27M mutations, which result in aberrant signaling, are identified. We evaluated the CNS exposure of the DNA methyltransferase inhibitor, 5-azacytidine (5-AZA), in preclinical nonhuman primate (NHP) models to inform its clinical development for CNS tumors. METHODS: 5-AZA and 5-AZA+Inulin pharmacokinetics (PK) were evaluated in NHPs (n = 10) following systemic (intravenous [IV]) and intrathecal (intraventricular [IT-V], intralumbar [IT-L], and cisternal [IT-C]) administration. Plasma, cerebrospinal fluid (CSF), cortical extracellular fluid (ECF), and tissues were collected. 5-AZA levels were quantified via ultra-high-performance liquid chromatography with tandem mass spectrometric detection assay and inulin via ELISA. PK parameters were calculated using noncompartmental methods. RESULTS: After IV administration, minimal plasma exposure (area under the curve [AUC] range: 2.4-3.2 h*µM) and negligible CSF exposure were noted. CSF exposure was notably higher after IT-V administration (AUCINF 1234.6-5368.4 h*µM) compared to IT-L administration (AUCINF 7.5-19.3 h*µM). CSF clearance after IT administration exceeded the mean inulin CSF flow rate of 0.018 ± 0.003 ml/min as determined by inulin IT-V administration. 5-AZA IT-V administration with inulin increased the 5-AZA CSF duration of exposure by 2.2-fold. IT-C administration yielded no quantifiable 5-AZA ECF concentrations but resulted in quantifiable tissue levels. CONCLUSIONS: IT administration of 5-AZA is necessary to achieve adequate CNS exposure. IT administration results in pronounced and prolonged 5-AZA CSF exposure above the reported IC50 range for IDH-mutated glioma cell lines. Inulin administered with 5-AZA increased the duration of exposure for 5-AZA.

11.
Comp Med ; 70(6): 526-531, 2020 12 01.
Artículo en Inglés | MEDLINE | ID: mdl-33046181

RESUMEN

Cerebrospinal fluid (CSF) flow rate and volume are fundamental to the design and interpretation of preclinical pharmacokinetics and pharmacodynamics studies in NHP. To determine the values of CSF flow rate and volume, we evaluated the plasma and CSF pharmacokinetics of inulin, an inert polysaccharide tracer, in 5 rhesus macaques with CSF ventricular res- ervoirs and lumbar ports; these reservoirs and ports facilitate humane intrathecal administration and serial CSF sampling in unanesthetized macaques. Inulin was administered intrathecally via the CSF ventricular reservoir (n = 3), followed by the collection of lumbar CSF via the lumbar port and plasma. The contribution of dietary inulin was evaluated by using pre- and postprandial inulin plasma concentrations (n = 2) and a feed analysis of the NHP diet. Inulin concentrations were quantified using ELISA. Pharmacokinetic parameters were calculated by using noncompartmental methods. Daily diet was analyzed for inulin by using Official Method no. 997.08 of AOAC International. In male rhesus macaques, the mean CSF flow rate, established via inulin clearance after IT administration, was 0.018 ± 0.003 mL/min; mean CSF volume, established based on apparent volume of distribution, was 10.17 ± 0.63 mL. In plasma, inulin was quantifiable in all pre-administration samples and increased over the sampling period, precluding interpretation of plasma pharmacokinetics. Evaluation of the effect of diet on plasma concentrations established quantifiable inulin levels that showed minimal variation relative to the prandial state. Analysis of the feed detected 5 inulin types ranging from 1100 to 1440 mg per100 g. The diet was the source of detectable pre-administration inulin plasma concentrations, whereas inulin was not detected in CSF before inulin administration.


Asunto(s)
Líquido Cefalorraquídeo , Inulina , Animales , Macaca mulatta , Masculino
12.
J Am Assoc Lab Anim Sci ; 58(2): 223-230, 2019 03 01.
Artículo en Inglés | MEDLINE | ID: mdl-30630557

RESUMEN

Regional anesthesia is a commonly used adjunct to orofacial dental and surgical procedures in companion animals and humans. However, appropriate techniques for anesthetizing branches of the mandibular and maxillary nerves have not been described for rhesus monkeys. Skulls of 3 adult rhesus monkeys were examined to identify relevant foramina, establish appropriate landmarks for injection, and estimate injection angles and depth. Cadaver heads of 7 adult rhesus monkeys (4 male, 3 female) were then injected with thiazine dye to demonstrate correct placement of solution to immerse specific branches of the mandibular and maxillary nerves. Different volumes of dye were injected on each side of each head to visualize area of diffusion, and to estimate the minimum volume needed to saturate the area of interest. After injection, the heads were dissected to expose the relevant nerves and skull foramina. We describe techniques for blocking the maxillary nerve as well as its branches: the greater palatine nerve, nasopalatine nerve, and infraorbital nerve. We also describe techniques for blocking branches of the mandibular nerve: inferior alveolar nerve, mental (or incisive) nerve, lingual nerve, and long buccal nerve. Local anesthesia for the mandibular and maxillary nerves can be accomplished in rhesus macaques and is a practical and efficient way to maximize animal welfare during potentially painful orofacial procedures.


Asunto(s)
Anestesia de Conducción/veterinaria , Odontología/veterinaria , Macaca mulatta/anatomía & histología , Procedimientos Quirúrgicos Operativos/veterinaria , Anestesia de Conducción/métodos , Animales , Cadáver , Femenino , Humanos , Nervio Lingual/anatomía & histología , Masculino , Mandíbula , Nervio Mandibular/anatomía & histología
13.
Sci Rep ; 9(1): 18965, 2019 12 12.
Artículo en Inglés | MEDLINE | ID: mdl-31831787

RESUMEN

Severe malarial anaemia (SMA) is the most common life-threatening complication of Plasmodium falciparum infection in African children. SMA is characterised by haemolysis and inadequate erythropoiesis, and is associated with dysregulated inflammatory responses and reduced complement regulatory protein levels (including CD35). However, a deeper mechanistic understanding of the pathogenesis requires improved animal models. In this comparative study of two closely related macaque species, we interrogated potential causal factors for their differential and temporal relationships to onset of SMA. We found that rhesus macaques inoculated with blood-stage Plasmodium coatneyi developed SMA within 2 weeks, with no other severe outcomes, whereas infected cynomolgus macaques experienced only mild/ moderate anaemia. The abrupt drop in haematocrit in rhesus was accompanied by consumption of haptoglobin (haemolysis) and poor reticulocyte production. Rhesus developed a greater inflammatory response than cynomolgus macaques, and had lower baseline levels of CD35 on red blood cells (RBCs) leading to a significant reduction in the proportion of CD35+ RBCs during infection. Overall, severe anaemia in rhesus macaques infected with P. coatneyi has similar features to SMA in children. Our comparisons are consistent with an association of low baseline CD35 levels on RBCs and of early inflammatory responses with the pathogenesis of SMA.


Asunto(s)
Anemia/sangre , Anemia/parasitología , Eritrocitos/metabolismo , Malaria/sangre , Plasmodium/metabolismo , Receptores de Complemento 3b/sangre , Anemia/patología , Animales , Eritrocitos/parasitología , Eritrocitos/patología , Femenino , Inflamación/sangre , Inflamación/parasitología , Inflamación/patología , Macaca fascicularis , Macaca mulatta , Malaria/parasitología , Malaria/patología , Especificidad de la Especie
14.
Nat Commun ; 10(1): 4300, 2019 09 20.
Artículo en Inglés | MEDLINE | ID: mdl-31541097

RESUMEN

Mainstay treatment for Plasmodium vivax malaria has long relied on chloroquine (CQ) against blood-stage parasites plus primaquine against dormant liver-stage forms (hypnozoites), however drug resistance confronts this regimen and threatens malaria control programs. Understanding the basis of P. vivax chloroquine resistance (CQR) will inform drug discovery and malaria control. Here we investigate the genetics of P. vivax CQR by a cross of parasites differing in drug response. Gametocytogenesis, mosquito infection, and progeny production are performed with mixed parasite populations in nonhuman primates, as methods for P. vivax cloning and in vitro cultivation remain unavailable. Linkage mapping of progeny surviving >15 mg/kg CQ identifies a 76 kb region in chromosome 1 including pvcrt, an ortholog of the Plasmodium falciparum CQR transporter gene. Transcriptional analysis supports upregulated pvcrt expression as a mechanism of CQR.


Asunto(s)
Antimaláricos/farmacología , Cloroquina/farmacología , Cruzamientos Genéticos , Resistencia a Medicamentos/genética , Proteínas de Transporte de Membrana/genética , Plasmodium vivax/efectos de los fármacos , Plasmodium vivax/genética , Proteínas Protozoarias/genética , Animales , Anopheles/parasitología , Culicidae/parasitología , Descubrimiento de Drogas , Femenino , Expresión Génica , Genes Protozoarios , Malaria/tratamiento farmacológico , Malaria Vivax/tratamiento farmacológico , Malaria Vivax/parasitología , Masculino , Plasmodium falciparum/genética
15.
Med Image Anal ; 46: 215-228, 2018 05.
Artículo en Inglés | MEDLINE | ID: mdl-29627686

RESUMEN

Recent improvements in cardiac computed tomography (CCT) allow for whole-heart functional studies to be acquired at low radiation dose (<2mSv) and high-temporal resolution (<100ms) in a single heart beat. Although the extraction of regional functional information from these images is of great clinical interest, there is a paucity of research into the quantification of regional function from CCT, contrasting with the large body of work in echocardiography and cardiac MR. Here we present the Simultaneous Subdivision Surface Registration (SiSSR) method: a fast, semi-automated image analysis pipeline for quantifying regional function from contrast-enhanced CCT. For each of thirteen adult male canines, we construct an anatomical reference mesh representing the left ventricular (LV) endocardium, obviating the need for a template mesh to be manually sculpted and initialized. We treat this generated mesh as a Loop subdivision surface, and adapt a technique previously described in the context of 3-D echocardiography to register these surfaces to the endocardium efficiently across all cardiac frames simultaneously. Although previous work performs the registration at a single resolution, we observe that subdivision surfaces naturally suggest a multiresolution approach, leading to faster convergence and avoiding local minima. We additionally make two notable changes to the cost function of the optimization, explicitly encouraging plausible biological motion and high mesh quality. Finally, we calculate an accepted functional metric for CCT from the registered surfaces, and compare our results to an alternate state-of-the-art CCT method.


Asunto(s)
Técnicas de Imagen Cardíaca/métodos , Interpretación de Imagen Radiográfica Asistida por Computador/métodos , Tomografía Computarizada por Rayos X/métodos , Algoritmos , Animales , Medios de Contraste , Perros , Yopamidol
16.
J Heart Lung Transplant ; 37(8): 967-975, 2018 08.
Artículo en Inglés | MEDLINE | ID: mdl-29933912

RESUMEN

BACKGROUND: Observational studies suggest that cell-free DNA (cfDNA) is a biomarker of tissue injury in a range of conditions including organ transplantation. However, the lack of model systems to study cfDNA and its relevance to tissue injury has limited the advancements in this field. We hypothesized that the predictable course of acute humoral xenograft rejection (AHXR) in organ transplants from genetically engineered donors provides an ideal system for assessing circulating cfDNA as a marker of tissue injury. METHODS: Genetically modified pig donor hearts were heterotopically transplanted into baboons (n = 7). Cell-free DNA was extracted from pre-transplant and post-transplant baboon plasma samples for shotgun sequencing. After alignment of sequence reads to pig and baboon reference sequences, we computed the percentage of xenograft-derived cfDNA (xdcfDNA) relative to recipient by counting uniquely aligned pig and baboon sequence reads. RESULTS: The xdcfDNA percentage was high early post-transplantation and decayed exponentially to low stable levels (baseline); the decay half-life was 3.0 days. Post-transplantation baseline xdcfDNA levels were higher for transplant recipients that subsequently developed graft loss than in the 1 animal that did not reject the graft (3.2% vs 0.5%). Elevations in xdcfDNA percentage coincided with increased troponin and clinical evidence of rejection. Importantly, elevations in xdcfDNA percentage preceded clinical signs of rejection or increases in troponin levels. CONCLUSION: Cross-species xdcfDNA kinetics in relation to acute rejection are similar to the patterns in human allografts. These observations in a xenotransplantation model support the body of evidence suggesting that circulating cfDNA is a marker of tissue injury.


Asunto(s)
Biomarcadores/sangre , Ácidos Nucleicos Libres de Células/sangre , Trasplante de Corazón/efectos adversos , Trasplante Heterólogo/efectos adversos , Enfermedad Aguda , Animales , Estudios de Seguimiento , Rechazo de Injerto/inmunología , Semivida , Organismos Modificados Genéticamente , Papio , Porcinos , Trasplante Heterotópico , Troponina/sangre
17.
Comp Med ; 57(1): 74-81, 2007 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-17348294

RESUMEN

Two natural outbreaks of mouse minute virus (MMV) are described. Observations during management of the naturally infected colonies led to a study in which 4-wk-old C57BL/6NCr and C57BL/6Tac mice were inoculated oronasally with an immunosuppressive variant of MMV (MMVi), as were adult C57BL/6NCr lactating dams or their pups (age, 10 d). By day 28 postinoculation, 100% of the 4-wk-old male C57BL/6NCr and C57BL/6Tac mice, 56.2% of 4-wk-old C57BL/6NCr female and 62.5% of 4-wk-old C57BL/6Tac female mice, 100% of adult lactating C57BL/6NCr dams, and 100% of inoculated pups (10 d) had seroconverted. Serologically positive nursing dams did not infect their nursing pups. In contrast, when nursing pups were inoculated, 100% of their dams seroconverted by 28 d postinoculation. Only 1 of 4 facility sentinels (Tac:SW female mice) seroconverted to MMVi and none of the 4 research sentinels (Tac:SW female mice) seroconverted under a once-weekly bedding transfer program. Consequently, 4 new research Tac:SW sentinels of each gender (n = 8) were placed in known-positive cages at cage-change; 100% of the male mice but 0% of the females seroconverted by day 48. Study results suggest gender influences both infectivity and the ability to detect subclinical infections of MMVi. Other factors that may influence detection of MMV include mouse strain or stock, short shedding period, and prolonged time between cage changes. In light of the data from both the natural infections and the experimental cases, cessation of breeding likely will be beneficial when trying to eradicate this virus.


Asunto(s)
Anticuerpos Antivirales/sangre , Ratones , Virus Diminuto del Ratón/inmunología , Infecciones por Parvoviridae/veterinaria , Enfermedades de los Roedores/inmunología , Enfermedades de los Roedores/virología , Animales , Cruzamiento/métodos , Femenino , Masculino , Ratones Endogámicos C57BL , Infecciones por Parvoviridae/inmunología , Reacción en Cadena de la Polimerasa , Vigilancia de Guardia/veterinaria , Factores Sexuales , Factores de Tiempo
18.
Int J Cardiovasc Imaging ; 33(8): 1253-1261, 2017 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-28289990

RESUMEN

To determine the feasibility of dual-contrast agent imaging of the heart using photon-counting detector (PCD) computed tomography (CT) to simultaneously assess both first-pass and late enhancement of the myocardium. An occlusion-reperfusion canine model of myocardial infarction was used. Gadolinium-based contrast was injected 10 min prior to PCD CT. Iodinated contrast was infused immediately prior to PCD CT, thus capturing late gadolinium enhancement as well as first-pass iodine enhancement. Gadolinium and iodine maps were calculated using a linear material decomposition technique and compared to single-energy (conventional) images. PCD images were compared to in vivo and ex vivo magnetic resonance imaging (MRI) and histology. For infarct versus remote myocardium, contrast-to-noise ratio (CNR) was maximal on late enhancement gadolinium maps (CNR 9.0 ± 0.8, 6.6 ± 0.7, and 0.4 ± 0.4, p < 0.001 for gadolinium maps, single-energy images, and iodine maps, respectively). For infarct versus blood pool, CNR was maximum for iodine maps (CNR 11.8 ± 1.3, 3.8 ± 1.0, and 1.3 ± 0.4, p < 0.001 for iodine maps, gadolinium maps, and single-energy images, respectively). Combined first-pass iodine and late gadolinium maps allowed quantitative separation of blood pool, scar, and remote myocardium. MRI and histology analysis confirmed accurate PCD CT delineation of scar. Simultaneous multi-contrast agent cardiac imaging is feasible with photon-counting detector CT. These initial proof-of-concept results may provide incentives to develop new k-edge contrast agents, to investigate possible interactions between multiple simultaneously administered contrast agents, and to ultimately bring them to clinical practice.


Asunto(s)
Medios de Contraste/administración & dosificación , Corazón/diagnóstico por imagen , Yopamidol/administración & dosificación , Meglumina/administración & dosificación , Infarto del Miocardio/diagnóstico por imagen , Daño por Reperfusión Miocárdica/diagnóstico por imagen , Compuestos Organometálicos/administración & dosificación , Fotones , Cloruro de Potasio/administración & dosificación , Tomografía Computarizada por Rayos X/métodos , Animales , Cicatriz/diagnóstico por imagen , Cicatriz/patología , Modelos Animales de Enfermedad , Perros , Estudios de Factibilidad , Interpretación de Imagen Asistida por Computador , Modelos Lineales , Imagen por Resonancia Magnética , Infarto del Miocardio/patología , Daño por Reperfusión Miocárdica/patología , Miocardio/patología , Valor Predictivo de las Pruebas , Prueba de Estudio Conceptual , Reproducibilidad de los Resultados , Relación Señal-Ruido , Factores de Tiempo
19.
Comp Med ; 66(4): 349-52, 2016.
Artículo en Inglés | MEDLINE | ID: mdl-27538866

RESUMEN

Biomedical translational research frequently incorporates collection of CSF from NHP, because CSF drug levels are used as a surrogate for CNS tissue penetration in pharmacokinetic and dynamic studies. Surgical placement of a CNS ventricular catheter reservoir for CSF collection is an intensive model to create and maintain and thus may not be feasible or practical for short-term studies. Furthermore, previous NHP lumbar port models require laminectomy for catheter placement. The new model uses a minimally invasive technique for percutaneous placement of a lumbar catheter to create a closed, subcutaneous system for effective, repeated CSF sample collection. None of the rhesus macaques (Macaca mulatta; n = 10) implanted with our minimally invasive lumbar port (MILP) system experienced neurologic deficits, postoperative infection of the surgical site, or skin erosion around the port throughout the 21.7-mo study. Functional MILP systems were maintained in 70% of the macaques, with multiple, high-quality, 0.5- to 1.0-mL samples of CSF collected for an average of 3 mo by using aspiration or gravitational flow. Among these macaques, 57% had continuous functionality for a mean of 19.2 mo; 50% of the cohort required surgical repair for port repositioning and replacement during the study. The MILP was unsuccessful in 2 macaques, at an average of 9.5 d after surgery. Nonpatency in these animals was attributed to the position of the lumbar catheter. The MILP system is an appropriate replacement for temporary catheterization and previous models requiring laminectomy and is a short-term alternative for ventricular CSF collection systems in NHP.


Asunto(s)
Catéteres de Permanencia/veterinaria , Macaca mulatta/líquido cefalorraquídeo , Animales , Cateterismo/métodos , Cateterismo/veterinaria , Vértebras Lumbares/cirugía , Masculino , Procedimientos Quirúrgicos Mínimamente Invasivos/métodos , Procedimientos Quirúrgicos Mínimamente Invasivos/veterinaria , Modelos Animales
20.
Contemp Top Lab Anim Sci ; 44(6): 8-14, 2005 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-16370573

RESUMEN

Dendrobatid frogs are studied primarily for the bioactive alkaloids found in their skin. Also known as poison-dart frogs, these animals accumulate toxic alkaloids from dietary sources. The function and uses of the many alkaloids, the alkaloid accumulation system, and the basic biology and physiology of the frogs themselves are of research interest. Here we overview the taxonomy of these frogs and some of the unique aspects of their natural biology and reproduction. We also describe the components of a successful laboratory housing system, including temperature, lighting, humidity, ventilation, nutrition, health considerations, and handling. A brief summary of dendrobatid research highlights is provided.


Asunto(s)
Crianza de Animales Domésticos/métodos , Fenómenos Fisiológicos Nutricionales de los Animales , Anuros/fisiología , Ambiente , Vivienda para Animales , Reproducción/fisiología , Animales , Anuros/genética , Femenino , Larva/crecimiento & desarrollo , Masculino
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