Macaque Skin Research Articles

Particulate matter 10 induces oxidative stress and apoptosis in rhesus macaques skin fibroblast.

Particulate matter (PM) is a major air pollutant that affects human health worldwide. PM can pass through the skin barrier, thus causing skin diseases such as heat rash, allergic reaction, infection, or inflammation. However, only a few studies have been conducted on the cytotoxic effects of PM exposure on large-scale animals. Therefore, herein, we investigated whether and how PM affects rhesus macaque skin fibroblasts. Rhesus macaque skin fibroblasts were treated with various concentrations of PM10 (1, 5, 10, 50, and 100 μg/mL) and incubated for 24, 48, and 72 h. Then, cell viability assay, TUNEL assay, and qRT-PCR were performed on the treated cells. Further, the reactive oxygen species, glutathione, and cathepsin B levels were determined. The MTT assay revealed that PM10 (>50 μg/mL) proportionately reduced the cell proliferation rate. PM10 treatment increased TUNEL-positive cell numbers, following the pro-apoptosis-associated genes (CASP3 and BAX) and tumor suppressor gene TP53 were significantly upregulated. PM10 treatment induced reactive oxidative stress. Cathepsin B intensity was increased, whereas GSH intensity was decreased. The mRNA expression levels of antioxidant enzyme-related genes (CAT, GPX1 and GPX3) were significantly upregulated. Furthermore, PM10 reduced the mitochondrial membrane potential. The mRNA expression of mitochondrial complex genes, such as NDUFA1, NDUFA2, NDUFAC2, NDUFS4, and ATP5H were also significantly upregulated. In conclusion, these results showed that PM10 triggers apoptosis and mitochondrial damage, thus inducing ROS accumulation. These findings provide potential information on the cytotoxic effects of PM10 treatment and help to understand the mechanism of air pollution-induced skin diseases.

Intradermal vaccination prevents anti-MOG autoimmune encephalomyelitis in macaques.

BackgroundAutoimmune demyelinating diseases (ADD) are a major cause of neurological disability due to autoreactive cellular and humoral immune responses against brain antigens. A cure for chronic ADD could be obtained by appropriate immunomodulation.MethodsWe implemented a preclinical scheme to foster immune tolerance to myelin oligodendrocyte glycoprotein (MOG), in a cynomolgus-macaque model of experimental autoimmune encephalomyelitis (EAE), in which administration of recombinant human MOG (rhMOG) elicits brain inflammation mediated by MOG-autoreactive CD4+ lymphocytes and anti-MOG IgG. For immunotherapy, we used a recombinant antibody (Ab) directed against the dendritic cell-asialoglycoprotein receptor (DC-ASGPR) fused either to MOG or a control antigen PSA (prostate-specific antigen).FindingsrhMOG and the anti-DC-ASGPR-MOG were respectively detected in CD1a+ DCs or CD163+ cells in the skin of macaques. Intradermal administration of anti-DC-ASGPR-MOG, but not control anti-DC-ASGPR-PSA, was protective against EAE. The treatment prevented the CD4+ T cell activation and proinflammatory cytokine production observed in controls. Moreover, the administration of anti-DC-ASGPR-MOG induced MOG-specific CD4+CD25+FOXP3+CD39+ regulatory lymphocytes and favoured an upsurge in systemic TGFβ and IL-8 upon rhMOG re-administration in vivo.InterpretationWe show that the delivery of an anti-DC-ASGPR-MOG allows antigen-specific adaptive immune modulation to prevent the breach of immune tolerance to MOG. Our findings pave the way for therapeutic vaccines for long-lasting remission to grave encephalomyelitis with identified autoantigens, such as ADD associated with anti-MOG autoantibodies.FundWork supported by the French ANR (ANR-11-INBS-0008 and ANR-10-EQPX-02-01), NIH (NIH 1 R01 AI 105066), the Baylor Scott and White Healthcare System funding and Roche Research Collaborative grants.

A biosafety evaluation of synchrotron radiation X-ray to skin and bone marrow: single dose irradiation study of rats and macaques

Purpose: Very limited experimental data is available regarding the safe dosages related to synchrotron radiation (SR) procedures. We used young rats and macaques to address bone marrow and skin tolerance to various doses of synchrotron radiation.Methods: Rats were subjected to 0, 0.5, 2.5, 5, 25 or 100 Gy local SR X-ray irradiation at left hind limb. Rat blood samples were analyzed at 2–90 days after irradiation. The SR X-ray irradiated skin and tibia were sectioned for morphological examination. For non-human primate study, three male macaques were subjected to 0.5 or 2.5 Gy SR X-ray on crus. Skin responses of macaques were observed.Results: All rats that received SR X-ray irradiation doses greater than 2.5 Gy experienced hair loss and bone-growth inhibition, which were accompanied by decreased number of follicles, thickened epidermal layer, and decreased density of bone marrow cells (p < 0.05). Macaque skin could tolerate 0.5 Gy SR X-ray but showed significant hair loss when the dose was raised above 2.5 Gy.Conclusion: The safety threshold doses of SR X-ray for rat skin, bone marrow and macaque skin are between 0.5 and 2.5 Gy. Our study provided essential information regarding the biosafety of SR X-ray irradiation.

Macrophage- and neutrophil-derived TNF-α instructs skin langerhans cells to prime antiviral immune responses.

Dendritic cells are major APCs that can efficiently prime immune responses. However, the roles of skin-resident Langerhans cells (LCs) in eliciting immune responses have not been fully understood. In this study, we demonstrate for the first time, to our knowledge, that LCs in cynomolgus macaque skin are capable of inducing antiviral-specific immune responses in vivo. Targeting HIV-Gag or influenza hemagglutinin Ags to skin LCs using recombinant fusion proteins of anti-Langerin Ab and Ags resulted in the induction of the viral Ag-specific responses. We further demonstrated that such Ag-specific immune responses elicited by skin LCs were greatly enhanced by TLR ligands, polyriboinosinic polyribocytidylic acid, and R848. These enhancements were not due to the direct actions of TLR ligands on LCs, but mainly dependent on TNF-α secreted from macrophages and neutrophils recruited to local tissues. Skin LC activation and migration out of the epidermis are associated with macrophage and neutrophil infiltration into the tissues. More importantly, blocking TNF-α abrogated the activation and migration of skin LCs. This study highlights that the cross-talk between innate immune cells in local tissues is an important component for the establishment of adaptive immunity. Understanding the importance of local immune networks will help us to design new and effective vaccines against microbial pathogens.

Epigenetic Reprogramming by Somatic Cell Nuclear Transfer in Primates

We recently demonstrated that somatic cells from adult primates could be reprogrammed into a pluripotent state by somatic cell nuclear transfer. However, the low efficiency with donor cells from one monkey necessitated the need for large oocyte numbers. Here, we demonstrate nearly threefold higher blastocyst development and embryonic stem (ES) cell derivation rates with different nuclear donor cells. Two ES cell lines were isolated using adult female rhesus macaque skin fibroblasts as nuclear donors and oocytes retrieved from one female, following a single controlled ovarian stimulation. In addition to routine pluripotency tests involving in vitro and in vivo differentiation into various somatic cell types, primate ES cells derived from reprogrammed somatic cells were also capable of contributing to cells expressing markers of germ cells. Moreover, imprinted gene expression, methylation, telomere length, and X-inactivation analyses were consistent with accurate and extensive epigenetic reprogramming of somatic cells by oocyte-specific factors.

Attenuated and wild-type HIV-1 infections and long terminal repeat-mediated gene expression from plasmids delivered by gene gun to human skin ex vivo and macaques in vivo.

Gene expression from HIV-based gene therapy vectors or live-attenuated HIV-1 vaccines requires RNA transcription supported by the HIV-1 promoter, the long terminal repeat (LTR). Delivery of live-attenuated HIV-1 vaccines as plasmid DNA would overcome problems associated with production of attenuated HIV-1 strains. We investigated the expression of reporter plasmids and proviral HIV-1 constructs driven by either the HIV-1 LTR or LTRs with deletions in the U3 enhancer regions. LTR-driven plasmids were inoculated by gene gun into both human epidermis ex vivo and macaques in vivo. The HIV-1 LTR drove reporter gene expression in human and macaque skin, although with 15- to 20-fold less efficiency compared to the immediate-early cytomegalovirus promoter. A deleted LTR derived from a naturally attenuated HIV-1 strain infecting a member of the well-characterized Sydney Blood Bank Cohort of long-term nonprogressors was 5-fold less efficient in expression of the reporter gene compared to wild-type LTR. Delivery of proviral wild-type HIV-1 DNA constructs to human skin resulted in recovery of HIV-1 from cells emigrating from the epidermis, providing an ex vivo model of the infectivity of proviral HIV-1 DNA. However, delivery of proviral HIV-1 DNA containing deletions in either the LTR, Nef, or the secondary viral transcription activator,Vpr, significantly reduced HIV-1 replication in this model. The early coexpression of Tat from a second plasmid did not restore replication. Thus, although attenuated lentiviral vaccines might be deliverable as proviral DNA constructs in primate subjects, significant improvements are needed to enhance the efficiency of this method.

Dendrite cell-T cell mixtures, isolated from the skin and mucosae of macaques, support the replication of SIV.

Previous studies have shown that HIV-1 exploits dendritic cells (DCs) to replicate and spread among CD4+ T cells. The DCs within mucosal surfaces may be especially important, but these are more difficult to access. To study more extensively the properties of DCs and other leukocytes from skin and different mucosae, DCs were isolated from uninfected macaques and their sensitivity assessed to infection with SIV in vitro. Dendritic cells and T cells readily emigrated from organ cultures of macaque skin, as described previously for humans. In addition, characteristic cells emigrated from explants of mucosae, both nasopharyngeal (adenoid and tonsil) and genital (vagina and cervix). The macaque DCs reacted with the monoclonals that are used to study human DCs, such as MAbs to CD40, CD86, CD83, and the p55 protein. When SIV was added to the DC-T cell mixtures from these different organs, extensive replication was observed in all but the cervical leukocytes. SIV replication occurred without the use mitogens, and with virus that had been grown in a cell line in the absence of mitogens and IL-2. Most of the newly synthesized viral protein is observed in syncytia. Therefore, mixtures of DCs and T cells isolated from mucosal surfaces served as a naturally permissive environment for SIV replication.

EFFECT OF TOPICAL TRETINOIN ON EPIDERMAL LANGERHANS’CELLS IN PHOTODAMAGED SKIN

Topical tretinoin has been successfully applied to treat photoaging; however, a decrease in the number of Langerhans' cells (LC) has been reported after its topical application in Macaque skin. A study was performed to evaluate the possible effect of topical tretinoin on the number of LC in human beings. Eight patients were studied. Topical tretinoin was applied in progressively increasing concentrations: 0.025% for 1 month, 0.05% for one month and 0.1% for 4 months. A skin biopsy from the malar area was taken before this therapy and 6 months later. To study LC, 4 mu frozen sectionswere stained with the anti-CD1 antibody. The number of CD1+ cells did not change when they were counted per unit of epidermal length, but they decreased when they were counted per unit of epidermal surface. These results indicate that topical tretinoin might damage epidermal Langerhans' cells, when it is applied for long periods of time; future studies are necessary to clarify this point.

Action of topical minoxidil in the bald stump-tailed macaque

The effect of topical minoxidil (5% and 2% solutions) on hair regrowth was studied in the frontal bald scalp of 18 adolescent and adult stump-tailed macaques (Macaca arctoides). Gross observation of the hairiness and folliculogram analysis of the skin biopsy specimens have shown that minoxidil induces the enlargement of vellus follicles to the size of middle to terminal follicles (regrowth of hair effect), minoxidil maintains the terminal follicles in the prebald scalp of periadolescent animals (prevention of baldness effect), enlarged follicles regress after minoxidil is withdrawn, and hair follicular growth is once again stimulated when treatment with minoxidil is reinstituted. Hair regrowth was more prominent in the early stage of baldness among younger macaques than in baldness of longer duration in older animals. An in vitro study of 3H thymidine uptake revealed that the hair follicles in minoxidil-treated macaque skin showed significant enhancement of deoxyribonucleic acid synthesis in the follicular and perifollicular cells but not in the epidermal keratinocytes. Furthermore, the uptake of 3H minoxidil and its conversion to minoxidil sulfate (the active metabolite producing vasodilation) was relatively higher in the hair follicles than in the epidermis and dermis. Serum concentration of minoxidil was fairly constant 2, 4, 6, 15, and 24 hours after a single application (averaging 15 ng/ml with 5% minoxidil). Minoxidil's essential action in hair follicular growth may be as a potent vasodilator. However, a direct action on the hair follicle cannot be ruled out considering uptake and conversion of the drug to minoxidil sulfate within the hair follicle itself.

Immunohistochemical demonstration of neuron specific enolase (NSE) on cutaneous nerves: Comparative study using NSE and 5-100 protein antibodies on denervated skin

The presence of neuron-specific enolase (NSE) and S-100 protein, which are nerve specific proteins, was immunohistochemically investigated on the cutaneous nerves. NSE was found in the axons of the cutaneous nerve bundles and the terminal axons in the Meissner and Pacinian corpuscles of the normal human and macaque skin. S-100 protein was found in the Schwann cells, lamellar cells of the Meissner corpuscles, and inner core cells of the Pacinian corpuscles. After denervation of the ulnar nerve on macaque, NSE on axons of the cutaneous nerves and Meissner and Pacinian corpuscles was completely disappeared in the 5th digit. However, S-100 protein was still maintained in the Schwann cells and Meissner and Pacinian corpuscles in the same digit. From these results, we conclude that the comparative immunohistochemical staining of NSE and S-100 protein is simple and reliable method to demonstrate the cutaneous nerves in normal and pathological conditions.