Single-nucleus transcriptomics reveals a gatekeeper role for FOXP1 in primate cardiac aging

Aging poses a major risk factor for cardiovascular diseases, the leading cause of death in the aged population. However, the cell type-specific changes underlying cardiac aging are far from being clear. Here, we performed single-nucleus RNA-sequencing analysis of left ventricles from young and aged cynomolgus monkeys to define cell composition changes and transcriptomic alterations across different cell types associated with age. We found that aged cardiomyocytes underwent a dramatic loss in cell numbers and profound fluctuations in transcriptional profiles. Via transcription regulatory network analysis, we identified FOXP1, a core transcription factor in organ development, as a key downregulated factor in aged cardiomyocytes, concomitant with the dysregulation of FOXP1 target genes associated with heart function and cardiac diseases. Consistently, the deficiency of FOXP1 led to hypertrophic and senescent phenotypes in human embryonic stem cell-derived cardiomyocytes. Altogether, our findings depict the cellular and molecular landscape of ventricular aging at the single-cell resolution, and identify drivers for primate cardiac aging and potential targets for intervention against cardiac aging and associated diseases.

Establishment of anterior cruciate ligament reconstruction model in cynomolgus monkey with autogenous hamstring tendon transplantation / 中国修复重建外科杂志

OBJECTIVE@#To investigate the feasibility of establishing an anterior cruciate ligament (ACL) reconstruction model using hamstring tendon autograft in cynomolgus monkeys.@*METHODS@#Twelve healthy adult male cynomolgus monkeys, weighing 8-13 kg, were randomly divided into two groups ( n=6). In the experimental group, the ACL reconstruction model of the right lower limb was prepared by using a single bundle of hamstring tendon, and the ACL of the right lower limb was only cut off in the control group. The survival of animals in the two groups was observed after operation. Before operation and at 3, 6, and 12 months after operation, the knee range of motion, thigh circumference, and calf circumference of the two groups were measured; the anterior tibial translation D-value (ATTD) was measured by Ligs joint ligament digital body examination instrument under the loads of 13-20 N, respectively. At the same time, the experimental group underwent MRI examination to observe the graft morphology and the signal/ noise quotient (SNQ) was caculated.@*RESULTS@#All animals survived to the end of the experiment. In the experimental group, the knee range of motion, thigh circumference, and calf circumference decreased first and then gradually increased after operation; the above indexes were significantly lower at 3 and 6 months after operation than before operation ( P<0.05), and no significant difference was found between pre-operation and 12 months after operation ( P>0.05). In the control group, there was no significant change in knee range of motion after operation, showing no significant difference between pre- and post-operation ( P>0.05), but the thigh circumference and calf circumference gradually significantly decreased with time ( P<0.05), and the difference was significant when compared with those before operation ( P<0.05). At 6 and 12 months after operation, the thigh circumference and calf circumference were significantly larger in the experimental group than in the control group ( P<0.05). At 3 and 6 months after operation, the knee range of motion was significantly smaller in the experimental group than in the control group ( P<0.05). Under the loading condition of 13-20 N, the ATTD in the experimental group increased first and then decreased after operation; and the ATTD significantly increased at 3, 6 months after operation when compared with the value before operation ( P<0.05). But there was no significant difference between the pre-operation and 12 months after operation ( P>0.05). There was no significant change in ATTD in the control group at 3, 6, and 12 months after operation ( P>0.05), and which were significantly higher than those before operation ( P<0.05). At each time point after operation, the ATTD was significantly smaller in the experimental group than in the control group under the same load ( P<0.05). The MRI examination of the experimental group showed that the ACL boundary gradually became clear after reconstruction and was covered by the synovial membrane. The SNQ at each time point after operation was significantly higher than that before operation, but gradually decreased with time, and the differences between time points were significant ( P<0.05).@*CONCLUSION@#The ACL reconstruction model in cynomolgus monkey with autogenous hamstring tendon transplantation was successfully established.

Techniques of orthotopic renal transplantation. II. Size-matched porcine grafts in monkey recipients

ABSTRACT Purpose As a classical xenotransplantation model, porcine kidneys have been transplanted into the lower abdomen of non-human primates. However, we have improved upon this model by using size-matched grafting in the orthotopic position. The beneficial aspects and surgical details of our method are reported herein. Methods Donors were two newborn pigs (weighting 5 to 6 kg) and recipients were two cynomolgus monkeys (weighting, approximately, 7 kg). After bilateral nephrectomy, kidneys were cold-transported in Euro-Collins solution. The porcine kidney was transplanted to the site of a left nephrectomy and fixed to the peritoneum. Results Kidneys transplanted to the lower abdomen by the conventional method were more susceptible to torsion of the renal vein (two cases). In contrast, early-stage blood flow insufficiency did not occur in orthotopic transplants of theleft kidney. Conclusions Size-matched porcine-primate renal grafting using our method of transplanting tothe natural position of the kidneys contributes to stable post-transplant blood flow to the kidney.

Generation of a Hutchinson-Gilford progeria syndrome monkey model by base editing

Many human genetic diseases, including Hutchinson-Gilford progeria syndrome (HGPS), are caused by single point mutations. HGPS is a rare disorder that causes premature aging and is usually caused by a de novo point mutation in the LMNA gene. Base editors (BEs) composed of a cytidine deaminase fused to CRISPR/Cas9 nickase are highly efficient at inducing C to T base conversions in a programmable manner and can be used to generate animal disease models with single amino-acid substitutions. Here, we generated the first HGPS monkey model by delivering a BE mRNA and guide RNA (gRNA) targeting the LMNA gene via microinjection into monkey zygotes. Five out of six newborn monkeys carried the mutation specifically at the target site. HGPS monkeys expressed the toxic form of lamin A, progerin, and recapitulated the typical HGPS phenotypes including growth retardation, bone alterations, and vascular abnormalities. Thus, this monkey model genetically and clinically mimics HGPS in humans, demonstrating that the BE system can efficiently and accurately generate patient-specific disease models in non-human primates.

Evaluation of fecal microbiomes associated with obesity in captive cynomolgus monkeys (Macaca fascicularis)

Microorganisms play important roles in obesity; however, the role of the gut microbiomes in obesity is controversial because of the inconsistent findings. This study investigated the gut microbiome communities in obese and lean groups of captive healthy cynomolgus monkeys reared under strict identical environmental conditions, including their diet. No significant differences in the relative abundance of Firmicutes, Bacteroidetes and Prevotella were observed between the obese and lean groups, but a significant difference in Spirochetes (p < 0.05) was noted. Microbial diversity and richness were similar, but highly variable results in microbial composition, diversity, and richness were observed in individuals, irrespective of their state of obesity. Distinct clustering between the groups was not observed by principal coordinate analysis using an unweighted pair group method. Higher sharedness values (95.81% ± 2.28% at the genus level, and 79.54% ± 5.88% at the species level) were identified among individual monkeys. This paper reports the association between the gut microbiome and obesity in captive non-human primate models reared under controlled environments. The relative proportion of Firmicutes and Bacteroidetes as well as the microbial diversity known to affect obesity were similar in the obese and lean groups of monkeys reared under identical conditions. Therefore, obesity-associated microbial changes reported previously appear to be associated directly with environmental factors, particularly diet, rather than obesity.

Comparative safety assessments of the biosimilar APZ001 and Erbitux in pre-clinical animal models

Abstract Purpose: To evaluate the toxicity of Erbitux as well as its biosimilar APZ001 antibody (APZ001) in pre-clinical animal models including mice, rabbits and cynomolgus monkeys. Methods: We performed analysis of normal behavior activity, autonomic and non-autonomic nervous functions, nervous-muscle functions, nervous excitability and sensorimotor functions on CD-1 mice. Subsequently, we studied that effects of APZ001 and Erbitux on respiratory system, cardiovascular system and kidney in Cynomolgus monkey models and performed local tolerance experiments on New Zealand rabbits. Results: The comparisons between APZ001 and Erbitux showed no significant differences in mice autonomic nervous system, nervous muscle functions, non-autonomic nervous functions, nervous excitability and sensorimotor functions between treated and untreated group (p>0.05). APZ001 and Erbitux showed negative effect on CD-1 mice in the present of pentobarbital sodium anesthesia (p>0.05). Single administrations of high, medium or low doses of APZ001 did not lead to monkey urine volume alterations (p>0.05). In human tissues, APZ001 and Erbitux showed positive signals in endocardium, lung type II alveolar epithelial cell and surrounding vessels, but showed negative results in kidney and liver tissues. No hemolysis phenomenon and serious side-effects in vessels and muscles were observed in rabbits when administrated with APZ001 and Erbitux respectively. Conclusion: The safety comparisons between APZ001 antibody and Erbitux showed that these two antibodies showed highly similarities in mice, rabbits and cynomolgus monkey animal models in consideration of pharmaceutical effects, indicating APZ001 might be a suitable substitute for Erbitux.

Technique for orthotopic liver transplantation in cynomolgus monkeys (Macaca fascicularis)

PURPOSE: Recent studies investigating new strategies to modulate the immune system have utilized animal models of liver transplantation (LT). However, the anhepatic phase (AHP) remains a crucial problem in LT. The aim of the present study is to introduce a technique for successful orthotopic LT in cynomolgus monkeys using an early-reperfusion strategy. METHODS: Orthotopicallo-LT was performed with seven donor/recipient pairs of cynomolgus monkeys. RESULTS: In 2 recipients, liver allografts were perfused after suprahepatic inferior vena cava (SHIVC), portal vein (PV), and infrahepatic inferior vena cava (IHIVC) anastomosis. To reduce the time of AHP in five recipients, liver allografts ware perfused after SHIVC and PV anastomosis while the IHIVC was not anastomosed. In the latter strategy, the AHP was reduced from 46 minutes to 31 minutes and a 24-hour survival rate of 80% was achieved. CONCLUSION: Our results indicate that an early-reperfusion strategy can be successfully used to establish a LT model in cynomolgus monkeys with a consistently high rate of animal survival.

Non-invasive Myocardial Strain Imaging to Evaluate Graft Failure in Cardiac Xenotransplantation / 대한이식학회지

BACKGROUND: The shortage of human hearts for allotransplantation makes xenotransplantation a possible option for controllable organ providers. To detect acute xenograft rejection, invasive biopsy seems inevitable; however, this occasionally results in poor incision wound healing or infection. To date, no method of noninvasive imaging for early detection of xenograft rejection has been established. We hypothesized that ultrasound speckle tracking would better detect xenograft failure than routine left ventricular ejection fractions (EF). METHODS: From August 2013 to July 2015, a total of six cardiac heterotopic xenotransplants (α 1, 3-galactosyltransferase gene-knockout porcine heart) into cynomolgus monkeys were monitored with echocardiography every 3 to 7 days. M-mode and two-dimensional (2D)-EF measurements and myocardial strain analyses were performed. Cardiac xenograft pathology was reviewed from the immediate postoperative biopsy, as well as the necropsy. RESULTS: Myocardial speckle tracking analysis was feasible in all six cases. The longest survival was 43 days. Only one pathology-proven immunologic rejection occurred. Cardiac xenograft failure appeared as two types: a dilated pattern with decreased EF or a myocardial-thickening pattern with preserved EF. Both antibody-mediated rejection (n=1) and sepsis-induced myocardial dysfunction (n=2) revealed decreased radial or circumferential strains, but normal-range EF. Xenograft functional decline was significant with respect to radial or circumferential strain (P=0.028), but not to conventional M-mode or 2D-EFs (P=0.600, P=0.340, respectively). CONCLUSIONS: Radial and circumferential strains were significantly decreased in both types of xenograft failure, regardless of EF. Further studies are warranted to correlate the strain analysis and immunopathological details.

Preclinical development of a humanized neutralizing antibody targeting HGF

Hepatocyte growth factor (HGF) and its receptor, cMET, play critical roles in cell proliferation, angiogenesis and invasion in a wide variety of cancers. We therefore examined the anti-tumor activity of the humanized monoclonal anti-HGF antibody, YYB-101, in nude mice bearing human glioblastoma xenografts as a single agent or in combination with temozolomide. HGF neutralization, The extracellular signal-related kinases 1 and 2 (ERK1/2) phosphorylation, and HGF-induced scattering were assessed in HGF-expressing cell lines treated with YYB-101. To support clinical development, we also evaluated the preclinical pharmacokinetics and toxicokinetics in cynomolgus monkeys, and human and cynomolgus monkey tissue was stained with YYB-101 to test tissue cross-reactivity. We found that YYB-101 inhibited cMET activation in vitro and suppressed tumor growth in the orthotopic mouse model of human glioblastoma. Combination treatment with YYB-101 and temozolomide decreased tumor growth and increased overall survival compared with the effects of either agent alone. Five cancer-related genes (TMEM119, FST, RSPO3, ROS1 and NBL1) were overexpressed in YYB-101-treated mice that showed tumor regrowth. In the tissue cross-reactivity assay, critical cross-reactivity was not observed. The terminal elimination half-life was 21.7 days. Taken together, the in vitro and in vivo data demonstrated the anti-tumor efficacy of YYB-101, which appeared to be mediated by blocking the HGF/cMET interaction. The preclinical pharmacokinetics, toxicokinetics and tissue cross-reactivity data support the clinical development of YYB-101 for advanced cancer.

Cynomolgus monkeys (Macaca fascicularis) experimentally infected with B19V and hepatitis A virus: no evidence of the co-infection as a cause of acute liver failure

This study was conducted to analyse the course and the outcome of the liver disease in the co-infected animals in order to evaluate a possible synergic effect of human parvovirus B19 (B19V) and hepatitis A virus (HAV) co-infection. Nine adult cynomolgus monkeys were inoculated with serum obtained from a fatal case of B19V infection and/or a faecal suspension of acute HAV. The presence of specific antibodies to HAV and B19V, liver enzyme levels, viraemia, haematological changes, and necroinflammatory liver lesions were used for monitoring the infections. Seroconversion was confirmed in all infected groups. A similar pattern of B19V infection to human disease was observed, which was characterised by high and persistent viraemia in association with reticulocytopenia and mild to moderate anaemia during the period of investigation (59 days). Additionally, the intranuclear inclusion bodies were observed in pro-erythroblast cell from an infected cynomolgus and B19V Ag in hepatocytes. The erythroid hypoplasia and decrease in lymphocyte counts were more evident in the co-infected group. The present results demonstrated, for the first time, the susceptibility of cynomolgus to B19V infection, but it did not show a worsening of liver histopathology in the co-infected group.

Reference values of clinical pathology parameters in cynomolgus monkeys (Macaca fascicularis) used in preclinical studies / 한국실험동물학회지

Nonhuman primates are increasingly used in biomedical research since they are highly homologous to humans compared to other rodent animals. However, there is limited reliable reference data of the clinical pathology parameters in cynomolgus monkeys, and in particular, only some coagulation and urinalysis parameters have been reported. Here, we reported the reference data of clinical chemical, hematological, blood coagulation, and urinalysis parameters in cynomolgus monkeys. The role of sex differences was analyzed and several parameters (including hematocrit, hemoglobin, red blood cell, blood urea nitrogen, total bilirubin, alkaline phosphatase, creatinine kinase, gamma-glutamyl tranferase, and lactate dehydrogenase) significantly differed between male and female subjects. In addition, compared to previous study results, lactate dehydrogenase, creatinine kinase, and aspartate aminotransferase showed significant variation. Interstudy differences could be affected by several factors, including age, sex, geographic origin, presence/absence of anesthetics, fasting state, and the analytical methods used. Therefore, it is important to deliberate with the overall reference indices. In conclusion, the current study provides a comprehensive and updated reference data of the clinical pathology parameters in cynomolgus monkeys and provides improved assessment criteria for evaluating preclinical studies or biomedical research.

Differential Cellular Tropism of Lentivirus and Adeno-Associated Virus in the Brain of Cynomolgus Monkey

Many researchers are using viruses to deliver genes of interest into the brains of laboratory animals. However, certain target brain cells are not easily infected by viruses. Moreover, the differential tropism of different viruses in monkey brain is not well established. We investigated the cellular tropism of lentivirus and adeno-associated virus (AAV) toward neuron and glia in the brain of cynomolgus monkeys (Macaca fascularis). Lentivirus and AAV were injected into putamen of the monkey brain. One month after injection, monkeys were sacrificed, and then the presence of viral infection by expression of reporter fluorescence proteins was examined. Tissues were sectioned and stained with NeuN and GFAP antibodies for identifying neuronal cells or astrocytes, respectively, and viral reporter GFP-expressing cells were counted. We found that while lentivirus infected mostly astrocytes, AAV infected neurons at a higher rate than astrocytes. Moreover, astrocytes showed reactiveness when cells were infected by virus, likely due to virus-mediated neuroinflammation. The Sholl analysis was done to compare the hypertrophy of infected and uninfected astrocytes by virus. The lentivirus infected astrocytes showed negligible hypertrophy whereas AAV infected astrocytes showed significant changes in morphology, compared to uninfected astrocytes. In the brain of cynomolgus monkey, lentivirus shows tropism for astrocytes over neurons without much reactivity in astrocytes, whereas AAV shows tropism for neurons over glial cells with a significant reactivity in astrocytes. We conclude that AAV is best-suited for gene delivery to neurons, whereas lentivirus is the best choice for gene delivery to astrocytes in the brain of cynomolgus monkeys.

Development and Evaluation of Acellular Dermal Matrix Originated from Transgenic Porcine Skin for Primates Skin Graft

This study was performed to compare the healing quality of the allogenic acellular dermal matix (ADM) and xenogenic ADM combined with autologous split thicknessskin graft. Xenogenic ADM was obtained from two wild type pigs. Allogenic ADM was obtained from cynomolgus monkeys. ADM was stored with cryo-preservation. Full-thickness skin wounds were made on the back of two cynomolgus monkeys. In one monkey, wounds were covered by allogenic ADM combined with autologous split thickness skin graft (STSG) or autologous STSGonly. In another monkey, wounds were covered by xenogenic ADM combined with autologous skin graft or autologous skin graft only. Skin healing process was observed during 2 weeks and skin biopsies were performed on 3 months after skin transplantation. We obtained IACUC approval (ORIENT-IACUC-16053). Skin on the xenogenic ADM was necrotized 1 week after skin transplantation. Possibly due to the thickness of ADM, which block the blood supply from the subcutaneous tissue to the autologous skin graft. Skin biopsy revealed that less fibrotic change of the skin on the ADM compared with the skin without ADM. Xenogenic ADM can be used in high degree burn patients who can suffered from contracture after healing since it can reduce fibrotic change.

Reference values of hematology, biochemistry, and blood type in cynomolgus monkeys from cambodia origin / 한국실험동물학회지

Cynomolgus monkeys as nonhuman primates are valuable animal models because they have a high level of human gene homology. There are many reference values for hematology and biochemistry of Cynomolgus monkeys that are needed for proper clinical diagnosis and biomedical research conduct. The body weight information and blood type are also key success factors in allogeneic or xenogeneic models. Moreover, the biological parameters could be different according to the origin of the Cynomolgus monkey. However, there are limited references provided, especially of Cambodia origin. In this study, we measured average body weight of 2,518 Cynomolgus monkeys and analyzed hematology and serum biochemistry using 119 males, and determined blood types in 642 monkeys with Cambodia origin. The average body weight of male Cynomolgus monkeys were 2.56±0.345 kg and female group was 2.43±0.330 kg at the age from 2 to 3 years. The male group showed relatively sharp increased average body weight from the 3 to 4 age period compared to the female group. In hematology and biochemistry, it was found that most of the data was similar when compared to other references even though some results showed differences. The ABO blood type result showed that type A, B, AB, and O was approximately 15.6, 33.3, 44.2, and 6.9%, respectively. The main blood type in this facility was B and AB. These biological background references of Cambodia origin could be used to provide important information to researchers who are using them in their biomedical research.

Dormancy models for Mycobacterium tuberculosis: A minireview

Dormancy models for Mycobacterium tuberculosis play important roles in understanding various aspects of tuberculosis pathogenesis and in the testing of novel therapeutic regimens. By simulating the latent tuberculosis infection, in which the bacteria exist in a non-replicative state, the models demonstrate reduced susceptibility to antimycobacterial agents. This minireview outlines the models available for simulating latent tuberculosis both in vitro and in several animal species. Additionally, this minireview discusses the advantages and disadvantages of these models for investigating the bacterial subpopulations and susceptibilities to sterilization by various antituberculosis drugs.

Using immunoglobulin Y as an alternative antibody for the detection of hepatitis A virus in frozen liver sections

An increasing amount of research has been conducted on immunoglobulin Y (IgY) because the use of IgY offers several advantages with respect to diagnostic testing, including its easy accessibility, low cost and translatability to large-scale production, in addition to the fact that it can be ethically produced. In a previous work, immunoglobulin was produced and purified from egg yolks (IgY) reactive to hepatitis A virus (HAV) antigens. In the present work, this anti-HAV-specific IgY was used in an indirect immunofluorescence assay to detect viral antigens in liver biopsies that were obtained from experimentally infected cynomolgus monkeys. Fields that were positive for HAV antigen were detected in liver sections using confocal microscopy. In conclusion, egg yolks from immunised hens may be a reliable source for antibody production, which can be employed for immunological studies.

Anticorpos IgY específicos para rotavírus do grupo A. Uma abordagem terapêutica para rotavirose em Macaca fascicularis / Specific IgY antibodies to rotavirus group A. A therapeutic approach for rotavirus disease in macaca fasciculares

A produção de anticorpos em aves imunizadas seguida da extração desses anticorpos da gema dos ovos (IgY), tem atraído o interesse da comunidade científica, como pode ser demonstrado pelo aumento significativo da literatura sobre a IgY. [...] Destaca-se que a tecnologia da IgY oferece novas possibilidades de aplicação em imunoterapia e métodos de diagnóstico, tanto para aplicação humana quanto veterinária, incluindo estratégias de tratamentode doenças intestinais graves em crianças, particularmente em países pobres. Neste presente estudo, objetivou-se avaliar a eficiência terapêutica da IgY utilizando macacos cynomolgus (Macaca fascicularis) jovens desafiados com o rotavírus do grupo A (RVA) humano, a maior causa de morbidade e mortalidade de crianças em todo o mundo, especialmente em países em desenvolvimento. Para esta proposta, anticorpos IgY específicos contra o RVA foram produzidos em aves, purificados por polietileno glicol, caracterizados por eletroforese em gel depoliacrilamida, western blotting e um teste de neutralização em cultura de células (MA-104). Este experimento preliminar rendeu uma suspensão altamente concentrada de IgY específica antirotavírus(IgY anti-RVA) (média de 37 mg/mL). O macaco cynomolgus foi estabelecido como modelo de infecção experimental após uma única administração de suspensão de rotavírus humano (3,1x106FFU/mL) por sonda gástrica. Os animais foram acompanhados durante onze dias, sendo observadas as manifestações clínicas, cargas virais sérica e fecal, hematologia e dosagem de eletrólitos séricos...

The production of antibodies in chickens and the extraction of specific antibody suspensions fromegg yolk (IgY) are increasingly attracting the interest of the scientific community, asdemonstrated by the significant growth of the IgY literature. [...] Of note, the IgY-technology offersnew possibilities for application in human and veterinary diagnostics and therapeutics, includingstrategies for the treatment of severe intestinal diseases in children, particularly in poor countries.In this study, we aimed to evaluate the therapeutic efficacy of the IgY by using youngcynomolgus monkeys (Macaca fascicularis) challenged with human rotavirus group A (RVA), amajor cause of morbidity and mortality in children worldwide, especially in developing countries.For this purpose, specific IgY antibodies against RVA were produced in hens, purified bypolyethylene glycol, characterized by polyacrylamide gel electrophoresis, western blotting and aneutralization assay in a cell culture system (MA-104). This preliminary experiment has yielded ahigh concentrated suspension of anti-rotavirus specific IgY (anti-RVA IgY) (average 37 mg/ml).The cynomolgus experimental infection model was established after a single administration of ahuman rotavirus suspension (3.1x106FFU/ml) by oral gavage. The confined animals werefollowed during a period of eleven days, observed for clinical signs, measurement of serum andfaecal viral load, and evaluation of hematology and serum electrolytes. The main clinical sign(observed in two of the seven inoculated monkeys) was diarrhea associated with a decrease inserum potassium during three days, followed by recovery...

Avaliação da aplicação terapêutica de imunoglobulina Y em macacos cynomolgus infectados com rotavírus humano / Evaluation of therapeutic application of immunoglobulin y in cynomolgus monkeys infected with rotavirus human

O rotavírus é a causa mais frequente de gastroenterite aguda (GA) em crianças menores decinco anos de idade em todo o mundo, sendo responsável por até 200 mil mortes anualmente.Sua disseminação ocorre pelo contato pessoa-pessoa, principalmente pela via de transmissãofecal-oral. Embora haja vacinas disponíveis e em desenvolvimento para a rotavirose, medidasalternativas são necessárias como, por exemplo, nos casos de surto. Dessa forma, a utilizaçãode imunoglobulina Y (IgY) em imunoterapia passiva é justificável. A IgY é a imunoglobulinaque predomina na circulação das aves, sendo transferida por secreção ativa do sangue para agema dos ovos, a partir da qual a purificação é realizada para obtenção dos anticorposespecíficos de interesse. As vantagens apresentadas por esta metodologia incluem: fácilobtenção, baixo custo e capacidade de produção em larga escala de modo adequado a umpadrão bioético mais atual. É crescente sua aplicação em métodos de diagnóstico eimunoterapia passiva. Neste estudo, avaliamos a infectividade do rotavírus A (RVA) humanoem macacos cynomolgus (Macaca fascicularis) e a aplicação terapêutica da IgY específicaanti-rotavírus após desafio com RVA pela via oral. Os animais que receberam o tratamentoforam divididos em dois grupos: um recebeu IgY apenas pela via oral e outro pela via oral eintravenosa. Os animais foram acompanhados por cinco dias e foram avaliados sinais clínicos,carga viral sérica e fecal, hematologia e dosagem de eletrólitos séricos. Além disso, buscou-sedefinir o perfil de células do sistema imune no sangue periférico, assim como detecção decitocinas no soro dos animais, ensaios de imunofluorescência para detecção da proteína nãoestrutural do rotavírus e também das células do sistema imune em cortes congelados deintestino...

Rotavirus is the most common cause of acute gastroenteritis in children under five years oldworldwide, accounting for about 200,000 deaths per year. Its spread is due to person to personcontact, mainly through fecal-oral transmission. Although there are vaccines available forrotavirus, alternative measures are required, for example, in outbreaks. Thus, the use ofimmunoglobulin Y (IgY) in passive immunotherapy is justified. The IgY is the predominantimmunoglobulin in birds circulation, being transferred from the blood by active secretion intothe yolk of eggs from which the purification is performed to obtain the specific antibody ofinterest. The advantages presented by this methodology include: easy obtainment, low cost,and production capacity in large-scale, appropriate considering current bioethical guiderlines.IgY is increasing employed in methods of diagnosis and passive immunotherapy. This studyevaluated the infectivity of human rotavirus A (RVA) in cynomolgus monkeys (Macacafascicularis) and therapeutic function of IgY after challenge with RVA orally. The animalswere divided in two groups: one that received IgY only by oral route and the other by oral andintravenous routes. We followed up the animals for five days through clinical manifestations,serum and fecal viral load, hematology and dosage of serum electrolytes. Moreover, weinvestigated the profile of immune cells in peripheral blood, detection of cytokines in theserum, immunofluorescence assays for the detection of rotavirus non-structural protein andimmune cells in the intestine. The absence of diarrhea episodes was considered a good signfor the clinical efficacy of IgY immunotherapy, however, viral RNA was found in the stool ofsome animals. The group treated with IgY orally and intravenously was the one in which wedid not detect viral genome in faeces. As for the cell populations in peripheral blood, it wasnot observed significant difference between groups...

Adenovirus vector-mediated short hairpin RNA targeting nuclear factor-κB suppresses proliferation of endometrial cells of Macaca fascicularis in vitro / 南方医科大学学报

<p><b>OBJECTIVE</b>To assess the effect of a high specific adenovirus vector-mediated shRNA targeting nuclear factor-κB (NF-κB) on cell proliferation of the endometrium of Macaca fascicularis.</p><p><b>METHODS</b>The adenoviral vector NF-κB-p65-shRNA and the empty vector were separately trasnfected in cultured endometrial cells of Macaca fascicularis. The changes in the expression of the target gene protein and apoptotic proteins, cell proliferation, and cell cycle distribution were observed after the transfection.</p><p><b>RESULTS</b>Compared with the control cells, infection of the endometrial cells with the NF-κB-p65-shRNA adenovirus significantly increased the expression levels of apoptotic proteins, promoted apoptosis of the endometrial cells, and reduced the cells in division?stage.</p><p><b>CONCLUSIONS</b>NF-κB-p65 shRNA adenovirus can effectively promote apoptosis of endometrial cells and inhibit the proliferation of endometrial cells of Macaca fascicularis.</p>

Effect of alemtuzumab on intestinal intraepithelial lymphocytes and intestinal barrier function in cynomolgus model / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Alemtuzumab has been used in organ transplantation and a variety of hematologic malignancies (especially for the treatment of B-cell chronic lymphocytic leukemia). However, serious infectious complications frequently occur after treatment. The reason for increased infections postalemtuzumab treatment is unknown at this stage. We explore the effect of alemtuzumab on intestinal intraepithelial lymphocytes (IELs) and intestinal barrier function in cynomolgus model to explain the reason of infection following alemtuzumab treatment.</p><p><b>METHODS</b>Twelve male cynomolguses were randomly assigned to either a treatment or control group. The treatment group received alemtuzumab (3 mg/kg, intravenous injection) while the control group received the same volume of physiological saline. Intestinal IELs were isolated from the control group and the treatment group (on day 9, 35, and 70 after treatment) for counting and flow cytometric analysis. Moreover, intestinal permeability was monitored by enzymatic spectrophotometric technique and enzyme-linked immunosorbent assay.</p><p><b>RESULTS</b>The numbers of IELs were decreased significantly on day 9 after treatment compared with the control group (0.35 ± 0.07 × 10 8 and 1.35 ± 0.09 × 10 8 , respectively; P < 0.05) and were not fully restored until day 70 after treatment. There were significant differences among four groups considering IELs subtypes. In addition, the proportion of apoptotic IELs after alemtuzumab treatment was significantly higher than in the control group (22.01 ± 3.67 and 6.01 ± 1.42, respectively; P < 0.05). Moreover, the concentration of D-lactate and endotoxin was also increased significantly on day 9 after treatment.</p><p><b>CONCLUSIONS</b>Alemtuzumab treatment depletes lymphocytes in the peripheral blood and intestine of cynomolgus model. The induction of apoptosis is an important mechanism of lymphocyte depletion after alemtuzumab treatment. Notably, intestinal barrier function may be disrupted after alemtuzumab treatment.</p>