Population Genetics of the Washington National Primate Research Center's (WaNPRC) Captive Pigtailed Macaque (Macaca nemestrina) Population

Pigtailed macaques (Macaca nemestrina) provide an important model for biomedical research on human disease and for studying the evolution of primate behavior. The genetic structure of captive populations of pigtailed macaques is not as well described as that of captive rhesus (M. mulatta) or cynomolgus (M. fascicularis) macaques. The Washington National Primate Research Center houses the largest captive colony of pigtailed macaques located in several different housing facilities. Based on genotypes of 18 microsatellite (short tandem repeat [STR]) loci, these pigtailed macaques are more genetically diverse than captive rhesus macaques and exhibit relatively low levels of inbreeding. Colony genetic management facilitates the maintenance of genetic variability without compromising production goals of a breeding facility. The periodic introduction of new founders from specific sources to separate housing facilities at different times influenced the colony's genetic structure over time and space markedly but did not alter its genetic diversity significantly. Changes in genetic structure over time were predominantly due to the inclusion of animals from the Yerkes National Primate Research Center in the original colony and after 2005. Strategies to equalize founder representation in the colony have maximized the representation of the founders’ genomes in the extant population. Were exchange of animals among the facilities increased, further differentiation could be avoided. The use of highly differentiated animals may confound interpretations of phenotypic differences due to the inflation of the genetic contribution to phenotypic variance of heritable traits. Am. J. Primatol. 74:1017‐1027, 2012. © 2012 Wiley Periodicals, Inc.     

Catalogue of sounds of the Pigtailed macaque (Macaca nemestrina)

A catalogue of sounds from a laboratory population of Pigtailed macaques has been developed from spectograms. Sounds are divided into four general classes: clear, harsh and high-pitched calls, and miscellaneous sounds. It appears that all parameters of the vocal signal are independently manipulatable. A plot of selected spectral characteristics demonstrated the uniqueness of individual signals. In some instances, different sounds are developed by rearrangement of specific fragments, or by acoustical activities performed on similar sounds. This latter phenomenon, which will be termed mosaicism , parallels aspects of language design. Lastly, comparison of spectrograms for Pigtailed macaques with those published for Rhesus macaques ( Macaca mulatta ) reveals similarities in structure.     

Metastatic Hepatocellular Carcinoma in a Juvenile Rhesus Macaque (Macaca mulatta)

Neoplasia in juvenile (younger than 5 y) rhesus macaques has been estimated to represent only approximately 1.4% of all occurrences of spontaneous neoplasia. Here we report an unusual case of a 3.75-y-old primiparous female rhesus macaque that was euthanized due to poor prognosis associated with progressive anemia, marked hepatomegaly, and radiographic evidence of metastatic neoplasia. Postmortem examination revealed an invasive, hemorrhagic hepatic mass that effaced approximately 70% of the liver parenchyma and had evidence of metastatic spread to multiple abdominal organs, the lungs, and the pituitary gland. Neoplastic polygonal cells lined large necrohemorrhagic cavities and exhibited marked anisocytosis and anisokaryosis, with frequent multinucleate cells. There was no desmoplasia associated with the primary neoplasm or metastases. Immunohistochemical studies revealed the neoplastic cells to be diffusely reactive with pancytokeratin, cytokeratin 7, and cytokeratin 8/18 antibodies and rarely reactive with carcinoembryonic antigen antibodies. The cells did not react with vimentin, S100, CD31, or factor VIII antibodies. Tumor morphology and immunophenotype led to the diagnosis of anaplastic hepatocellular carcinoma. This report represents the first known case of metastatic liver neoplasia in a rhesus macaque. The young age of this animal and the aggressive nature of the neoplasm are highly unusual and reminiscent of adolescent onset hepatocellular carcinoma in humans.Abbreviations: CK, cytokeratin; HCC, Hepatocellular carcinomaNeoplasia in juvenile rhesus macaques (Macaca mulatta) is extremely uncommon.^17,18 In a recent review of spontaneous neoplasia in 2 colonies of rhesus macaques, animals younger than 5 y represented only 1.4% of the total number of cases, and primary hepatic tumors were uncommon.¹⁸ All 5 of the cases detected among 2660 macaques involved animals between 14 and 26.8 y of age, and none had evidence of metastasis. Primary hepatic tumors appear to be similarly infrequent occurrences in other nonhuman primates, with the notable exception of prosimians, in which tumors arising from the liver are common spontaneous neoplasms.^3,15,17,18 Malignant liver tumors account for only 1% of pediatric tumors in humans.^5,7,12 Approximately 80% of these are hepatoblastomas—neoplasms arising from liver progenitor cells—and hepatocellular carcinoma (HCC) represents the second most frequent diagnosis.^5,7,12The medicine department of the California National Primate Research Center evaluated a 3.75-y-old, sexually mature, female rhesus macaque for rapidly progressive disease associated with a hepatic mass. Anaplastic HCC with extensive metastasis was diagnosed after postmortem examination. The current report describes the clinical progression of disease, the gross and microscopic pathology of the affected macaque, and the immunohistochemical characterization of the neoplasm.     

Simian Varicella Virus in Pigtailed Macaques (Macaca nemestrina): Clinical, Pathologic, and Virologic Features

Simian varicella virus (SVV; Cercopithecine herpesvirus 9) is a naturally occurring herpesvirus of nonhuman primates. Here we present the clinical, pathologic, and virologic findings from 2 cases of SVV in adult female pigtailed macaques (Macaca nemestrina). The initial case presented with hyperthermia and a diffuse inguinal rash which spread centripetally, progressing to vesiculoulcerative dermatitis of the trunk, face, and extremities. At 96 h after presentation, the animal was anorexic and lethargic and had oral and glossal ulcerations. Euthanasia was elected in light of the macaque''s failure to respond to clinical treatment. Seven days after the first case was identified, a second macaque presented with a vesicular rash and was euthanized. Gross necropsy lesions for both cases included vesicular, ulcerative dermatitis with mucocutaneous extension and hepatic necrosis; the initial case also demonstrated necrohemorrhagic gastroenterocolitis and multifocal splenic necrosis. Histology confirmed herpetic viral infection with abundant intranuclear inclusion bodies. Immunofluorescence assays detected antibodies specific for SVV. PCR assays of vesicular fluid, tissue, and blood confirmed SVV and excluded varicella–zoster virus (Human herpesvirus 3). Serology for Macacine herpesvirus 1 (formerly Cercopithecine herpesvirus 1), poxvirus (monkeypox), and rubella was negative. Banked serum samples confirmed SVV exposure and seroconversion. Investigation into the epidemiology of the seroconversion demonstrated a SVV colony prevalence of 20%. The described cases occurred in animals with reconstituted immune systems (after total-body irradiation) and demonstrate the clinical effects of infection with an endemic infectious agent in animals with a questionable immune status.Abbreviations: IFA, immunofluorescence assay; SVV, simian varicella virus; TBI, total body irradiation; WaNPRC, Washington National Primate Research Center; VZV, varicella–zoster virus; McHv1, Macacine herpesvuris 1; SRV-2, Simian retrovirus 2 (type D)Simian varicella virus (SVV; Cercopithecine herpesvirus 9) is a naturally occurring herpesvirus of Old World primates responsible for sporadic epizootics in biomedical research facilities.² Signs of infection include fever, vesicular skin lesions, hemorrhagic ulceration throughout the gastrointestinal tract, and multifocal hemorrhagic necrosis of the liver, spleen, lymph nodes, and endocrine organs.^6,7,8 Other names for SVV include Delta herpesvirus, Liverpool vervet virus, patas herpesvirus, and Medical Lake macaque virus.^{16, 20-23} Like many other herpesviruses, SVV establishes persistent lifelong infections, with viral DNA detectable in neural ganglia.¹² Infection with SVV does not necessarily lead to lifelong latency, and periodic reactivation of SVV may occur.³ SVV is genetically and antigenically similar to Human herpesvirus 3,² commonly known as varicela–zoster virus (VZV), the etiologic agent of chickenpox and shingles in humans. SVV in macaques and VZV in man present with similar clinical signs; SVV has been proposed as an animal model of VZV disease in man.²⁴ Rarely, VZV may occur in higher primates (Gorilla).¹⁸ The 2 viruses must be distinguished from one another through molecular techniques.^1,410,11 Both viral infections are usually mild and self-limiting in immunocompetent hosts,^4,8 reactivation and viral shedding may occur during times of stress or immunosupression.^80,21,22A recent review of SVV in Old World Monkeys⁸ focused on SVV as a disease of nonhuman primates. This case report expands on the 2 most recent cases of SVV mentioned in that review.⁸ The animals described were housed in accordance with the regulations of the Animal Welfare Act and the recommendations of the Guide for the Care and Use of Laboratory Animals¹¹ at the Washington National Primate Research Center (WaNPRC) facility in Seattle. The Institutional Animal Care and Use Committee of the University of Washington approved all aspects of the study to which the animals were assigned. The 2 clinical cases described in this report originated at the WaNPR–Seattle facility; contact animals described originated at the WaNPR–Tulane facility. When animals are relocated between the 2 facilities, they are processed through a domestic quarantine consisting of isolation for 30 d, during which time 3 tuberculin skin tests, 2 physical examinations, and 1 complete blood count and serological panel are performed. The WaNPRC–Tulane facility houses a breeding colony founded by animals relocated to Louisiana from the WaNPRC–Medical Lake facility in 1996.     

Body Size Effects on the Acoustic Structure of Pigtail Macaque (Macaca nemestrina) Screams

Scream vocalizations produced by pigtail macaques during agonistic encounters were classified according to caller body weight (small, medium, large) on the basis of six frequency-related acoustic variables using direct discriminant analysis. Separate discriminant analyses were run for: 1. calls produced when the attack involved an opponent higher-ranking than the victim and contact occurred, and 2. calls produced when the opponent was higher-ranking, but no contact occurred. Screams were correctly classified as to the caller's weight class at levels significantly above those expected by chance alone. Screams given during contact aggression were classified significantly better than those given in the absence of contact. The effect of greater arousal level (fear) on the frequency range of calls may account for this difference. Macaque screams are representational signals that are important in the solicitation of agonistic aid from allies in the social group. That a relationship between body size and the frequency of screams is nonetheless evident argues for the fundamental nature of the relationship.     

Pneumatocele in a pig-tailed macaque (Macaca nemestrina)

Radiographic examination of a pig-tailed macaque (Macaca nemestrina) with pneumonia revealed a large pneumatocele. The pneumatocele, a thin-walled, partially fluid filled radiolucent area, occupied approximately one-third of the left thorax. Rapid resolution of the pneumatocele accompanied antimicrobial treatment of the pneumonia and coincided with clinical improvement. Severe pulmonary acariasis was found at postmortem 15 months later.     

Mineralized Trichobezoars in a Rhesus Macaque (Macaca mulatta)

A five-year-old female rhesus macaque (Macaca mulatta) presented with a thin body condition and multiple palpable mid-abdominal masses. Mineralized cecal trichobezoars were removed surgically. Thirteen months later, similar masses recurred and were confirmed with radiographs. This is the first case report of a mineralized cecal trichobezoar in a rhesus macaque.     

影响口腔鳞状细胞癌患者预后的临床病理因素分析

目的：研究影响口腔鳞状细胞癌（Oral squamous cellcar cinoma,OSCC）术后患者预后的临床病理因素。方法：回顾性研究55例手术治疗的原发口腔鳞状细胞癌患者与预后相关的因素：年龄、性别、发病部位、颈部淋巴结转移情况、肿瘤细胞分化程度等。结果：平均发病年龄为57.35±12.02岁,牙龈鳞癌的复发转移率最高（45.5%）,舌部第二（44%）,颊部第三（37.5%）,唇癌预后最好（0.0%）。术前颈部淋巴结转移情况、肿瘤细胞分化程度与预后有关。肿瘤细胞的分化程度与术前淋巴结转移无显著相关性。术前有颈部淋巴结转移合并中低分化与预后差相关。结论：口腔鳞状细胞癌的预后与发病部位无显著相关性。肿瘤中低分化及术前有淋巴结转移者易出现术后复发转移。     

影响口腔鳞状细胞癌患者预后的临床病理因素分析

目的:研究影响口腔鳞状细胞癌(Oral squamous cellcar cinoma,OSCC)术后患者预后的临床病理因素。方法:回顾性研究55例手术治疗的原发口腔鳞状细胞癌患者与预后相关的因素:年龄、性别、发病部位、颈部淋巴结转移情况、肿瘤细胞分化程度等。结果:平均发病年龄为57.35±12.02岁,牙龈鳞癌的复发转移率最高(45.5%),舌部第二(44%),颊部第三(37.5%),唇癌预后最好(0.0%)。术前颈部淋巴结转移情况、肿瘤细胞分化程度与预后有关。肿瘤细胞的分化程度与术前淋巴结转移无显著相关性。术前有颈部淋巴结转移合并中低分化与预后差相关。结论:口腔鳞状细胞癌的预后与发病部位无显著相关性。肿瘤中低分化及术前有淋巴结转移者易出现术后复发转移。     

CD9在口腔鳞状细胞癌中的表达水平及其临床意义

目的:口腔鳞状细胞癌是一类极易发生局部侵袭和淋巴结转移的恶性肿瘤,CD9蛋白在多种肿瘤的发生发展及侵袭转移过程中起到重要作用,本研究旨在分析CD9蛋白在口腔鳞状细胞癌中的表达水平及其临床意义。方法:收集我院诊断明确的口腔鳞癌肿瘤患者石蜡标本合计80例,通过免疫组化手段对CD9蛋白表达水平进行评价,并根据CD9蛋白的表达水平分组,分析患者的临床病理学特征与CD9蛋白的关系。结果:CD9在正常组织和癌旁组织正常表达,在肿瘤组织中表达率低,其表达水平和口腔鳞癌的分化程度,淋巴结转移及最终分期有相关性(P0.05)。结论:本研究结果揭示,CD9在口腔鳞状细胞癌的发生发展中起到重要作用,CD9蛋白水平的低表达或不表达可能预测着肿瘤具有更明显的恶性生物学行为,并可能成为口腔鳞状细胞癌预后的生物学指标及基因治疗的新靶点。     

VEGF-C 在口腔鳞状细胞癌中的表达及其意义

目的:检测血管内皮生长因子-C(VEGF-C)在口腔鳞状细胞癌(鳞癌)和癌旁组织中的表达情况,探讨VEGF-C在口腔鳞癌的增殖、浸润和淋巴转移中的作用。方法:采用免疫组化方法检测60例口腔鳞癌病人癌组织和癌旁组织中VEGF-C的表达,应用图像分析系统进行分析,用Spearman相关分析研究其与病变部位、肿瘤大小、病理分级、临床分期及颈淋巴结转移之间的关系。结果:口腔鳞癌组织VEGF-C的表达明显高于癌旁组织(u=7.747,P<0.01),其表达强度与临床分期及淋巴结转移密切相关(r=0.564、0.706,P<0.05),与病变部位、大小、病理分级无关。结论:口腔鳞癌细胞分泌VEGF-C诱导癌周淋巴管增生扩张是发生区域淋巴结转移的重要因素之一,VEGF-C有望作为早期临床判断和预测颈淋巴结转移的指标之一。     

Natural Infection of Burkholderia pseudomallei in an Imported Pigtail Macaque (Macaca nemestrina) and Management of the Exposed Colony

Identification of the select agent Burkholderia pseudomallei in macaques imported into the United States is rare. A purpose-bred, 4.5-y-old pigtail macaque (Macaca nemestrina) imported from Southeast Asia was received from a commercial vendor at our facility in March 2012. After the initial acclimation period of 5 to 7 d, physical examination of the macaque revealed a subcutaneous abscess that surrounded the right stifle joint. The wound was treated and resolved over 3 mo. In August 2012, 2 mo after the stifle joint wound resolved, the macaque exhibited neurologic clinical signs. Postmortem microbiologic analysis revealed that the macaque was infected with B. pseudomallei. This case report describes the clinical evaluation of a B. pseudomallei-infected macaque, management and care of the potentially exposed colony of animals, and protocols established for the animal care staff that worked with the infected macaque and potentially exposed colony. This article also provides relevant information on addressing matters related to regulatory issues and risk management of potentially exposed animals and animal care staff.Abbreviations: CDC, Centers for Disease Control and Prevention; IHA, indirect hemagglutination assay; PEP, postexposure prophylacticBurkholderia pseudomallei, formerly known as Pseudomonas pseudomallei, is a gram-negative, aerobic, bipolar, motile, rod-shaped bacterium. B. pseudomallei infections (melioidosis) can be severe and even fatal in both humans and animals. This environmental saprophyte is endemic to Southeast Asia and northern Australia, but it has also been found in other tropical and subtropical areas of the world.^7,22,32,42 The bacterium is usually found in soil and water in endemic areas and is transmitted to humans and animals primarily through percutaneous inoculation, ingestion, or inhalation of a contaminated source.^{8, 22,28,32,42} Human-to-human, animal-to-animal, and animal-to-human spread are rare.^8,32 In December 2012, the National Select Agent Registry designated B. pseudomallei as a Tier 1 overlap select agent.³⁹ Organisms classified as Tier 1 agents present the highest risk of deliberate misuse, with the most significant potential for mass casualties or devastating effects to the economy, critical infrastructure, or public confidence. Select agents with this status have the potential to pose a severe threat to human and animal health or safety or the ability to be used as a biologic weapon.³⁹Melioidosis in humans can be challenging to diagnose and treat because the organism can remain latent for years and is resistant to many antibiotics.^12,37,41 B. pseudomallei can survive in phagocytic cells, a phenomenon that may be associated with latent infections.^19,38 The incubation period in naturally infected animals ranges from 1 d to many years, but symptoms typically appear 2 to 4 wk after exposure.^13,17,35,38 Disease generally presents in 1 of 2 forms: localized infection or septicemia.²² Multiple methods are used to diagnose melioidosis, including immunofluorescence, serology, and PCR analysis, but isolation of the bacteria from blood, urine, sputum, throat swabs, abscesses, skin, or tissue lesions remains the ‘gold standard.’^9,22,40,42 The prognosis varies based on presentation, time to diagnosis, initiation of appropriate antimicrobial treatment, and underlying comorbidities.^7,28,42 Currently, there is no licensed vaccine to prevent melioidosis.There are several published reports of naturally occurring melioidosis in a variety of nonhuman primates (NHP; 2,10,13,17,25,30,31,35 The first reported case of melioidosis in monkeys was recorded in 1932, and the first published case in a macaque species was in 1966.³⁰ In the United States, there have only been 7 documented cases of NHP with B. pseudomallei infection.^2,13,17 All of these cases occurred prior to the classification of B. pseudomallei as a select agent. Clinical signs in NHP range from subclinical or subacute illness to acute septicemia, localized infection, and chronic infection. NHP with melioidosis can be asymptomatic or exhibit clinical signs such as anorexia, wasting, purulent drainage, subcutaneous abscesses, and other soft tissue lesions. Lymphadenitis, lameness, osteomyelitis, paralysis and other CNS signs have also been reported.^{2,7,10,22,28,32} In comparison, human''s clinical signs range from abscesses, skin ulceration, fever, headache, joint pain, and muscle tenderness to abdominal pain, anorexia, respiratory distress, seizures, and septicemia.^7,9,21,22

Table 1.

Summary of reported cases of naturally occurring Burkholderia pseudomalleiinfections in nonhuman primates

Polyostotic fibrous dysplasia in a cynomolgus Macaque (Macaca fascicularis)

A 2.3-y-old female cynomolgus macaque (Macaca fascicularis) presented with a broken right tibia and fibula. Radiographs showed multiple cyst-like defects in all long bones. We suspected that both fractures were pathologic because they occurred through these defects. Ultrasonography, MRI, and dual X-ray absorptiometry revealed that the defects were filled with soft tissue. Grossly, the bones were abnormal in shape, and a gelatinous material filled the defects and the surrounding marrow cavity. Histologically, the gelatinous material was composed of fibrin and cartilage; few normal bone cells were seen. Genetic testing revealed extra material on the short arm of chromosome 8 in all tissues examined, but no copy number alterations of likely clinical significance were observed, and no abnormalities were found that were unique to the lesions. In light of the clinical signs and radiographic and pathologic findings, polyostotic fibrous dysplasia was diagnosed. This report represents the first documented case of fibrous dysplasia in a cynomolgus macaque.     

Atrial Septal Defect in a Bonnet Macaque (Macaca radiata)

Atrial Septal Defect was detected at autopsy in a subadult bonnet macaque (Macaca radiata). Case history and autopsy findings were described.     

Population organization of wild pig-tailed macaques (Macaca nemestrina nemestrina) in West Sumatra

A field study on wild pig-tailed macaques was conducted in West Sumatra, Indonesia, during three periods from January 1985 to February 1987. During the nine months of the first two periods, unprovisioned monkeys were traced and observed. During the eight months of the last period, monkeys were provisioned and observed mainly at baiting sites. Three troops and ten solitary males appeared at the two baiting sites. Some males immigrated into and emigrated from the troops. The troops had a multi-male multi-female composition. The size of the various troops was 74, 49, and 81 individuals, respectively, and the mean adult sex ratio in the troops was 1:6.3; that is, markedly biased towards females. The home ranges of two of the troops overlapped considerably. When the troops encountered each other at the baiting sites, a clear dominance relationship was recognized. The troops differed in their integration as ranging units: two of the troops did not form subgroups (temporary fission and fusion of each troop), while the other troop frequently split into subgroups. Recent field studies on pig-tailed macaques have suggested a multi-leveled society with harem-type unit groups. However, in the present study, the troops observed had neither a substructure similar to harem-type groups nor a superstructure that emerged as a result of fusion of the troops. The unit group of the pig-tailed macaques appears to be a multi-male, matrilineal group.     

FOXP3 Subcellular Localization Predicts Recurrence in Oral Squamous Cell Carcinoma

Forkhead box protein P3 (FOXP3) expression in tumor infiltrating CD4⁺T cells is generally associated with an intrinsic capacity to suppress tumor immunity. Based on this notion, different studies have evaluated the prognostic value of this maker in cancer but contradictory results have been found. Indeed, even within the same cancer population, the presence of CD4⁺FOXP3⁺T cells has been associated,with either a poor or a good prognosis, or no correlation has beenfound. Here, we demonstrate,in patients with oral squamous cell carcinoma (OSCC), that what really represents a prognostic parameter is not the overall expression of FOXP3 but its intracellular localization.While overallFOXP3 expression in tumor infiltrating CD4⁺T cells does not correlate with tumor recurrence, its intracellular localization within the CD4 cells does: nuclear FOXP3 (nFOXP3) is associated with tumor recurrence within 3 years, while cytoplasmicFOXP3 (cFOXP3) is associated with a lower likelihood of recurrence. Thus, we propose elevated levels of the cFOXP3/nFOXP3 ratio within tumor infiltrating CD4⁺ T cells as a predictor of OSCC recurrence.     

VEGF在口腔鳞状细胞癌中的表达及其临床意义

目的:研究血管内皮生长因子(Vascular endothelial growth factor,VEGF)在口腔鳞状细胞癌中的表达及其临床意义.方法:免疫组化EnVisionTM法检测VEGF在57例手术治疗的原发性OSCC(口腔鳞状细胞癌)中的表迭.结果:①VEGF在OSCC中的阳性表达率为40.35%.②VEGF在不同发病部位组的表达:牙龈癌>舌癌>颊癌>唇癌,组间无统计学差异(P>0.05).③VEOF在不同病理分级组、不同预后组、不同年龄组、不同性别组的表达无显著性差异(P>0.05);淋巴结转移组的表达显著高于无转移组(P<0.05).结论:虽然VEGF的高表达可能在淋巴结转移中起一定的作用,但还不能作为预测口腔鳞状细胞癌临床生物学行为及预后的可靠指标.     

Agrin and Perlecan Mediate Tumorigenic Processes in Oral Squamous Cell Carcinoma

Oral squamous cell carcinoma is the most common type of cancer in the oral cavity, representing more than 90% of all oral cancers. The characterization of altered molecules in oral cancer is essential to understand molecular mechanisms underlying tumor progression as well as to contribute to cancer biomarker and therapeutic target discovery. Proteoglycans are key molecular effectors of cell surface and pericellular microenvironments, performing multiple functions in cancer. Two of the major basement membrane proteoglycans, agrin and perlecan, were investigated in this study regarding their role in oral cancer. Using real time quantitative PCR (qRT-PCR), we showed that agrin and perlecan are highly expressed in oral squamous cell carcinoma. Interestingly, cell lines originated from distinct sites showed different expression of agrin and perlecan. Enzymatically targeting chondroitin sulfate modification by chondroitinase, oral squamous carcinoma cell line had a reduced ability to adhere to extracellular matrix proteins and increased sensibility to cisplatin. Additionally, knockdown of agrin and perlecan promoted a decrease on cell migration and adhesion, and on resistance of cells to cisplatin. Our study showed, for the first time, a negative regulation on oral cancer-associated events by either targeting chondroitin sulfate content or agrin and perlecan levels.