Popliteal Node Research Articles

Sensitivity comparison for the Leishmania spp. detection in different canine tissues using PCR-HRM

Leishmaniasis is a vector-borne disease caused by a parasite protozoon from the genus Leishmania. Among the molecular techniques applied for detecting these parasites, real-time PCR with High Resolution Melting (PCR-HRM) proved advantageous since it simultaneously determines both the presence and species of the pathogen in one step, through amplification and later analysis of curves generated by melting temperature. Based on this molecular technique, the goal of this study was to estimate the PCR-HRM sensitivity for Leishmania spp. detection in different canine tissues by evaluating biological samples obtained from popliteal, submandibular, and pre-scapular lymph nodes, from bone marrow and ear pinnae of 28 stray dogs captured in the metropolitan area of Asunción (Paraguay). The rk39 immunochromatographic test showed that 25/28 tested dogs (89%) presented antibodies against L. infantum. In 20/25 dogs that tested positive for rk39 (80%), it was possible to detect Leishmania spp. by PCR-HRM and determine that the species corresponded entirely to L. infantum. Regarding the analysis of different tissues, the parasite was detected in all popliteal lymph node samples, followed by high detection in submandibular (at 95%) and pre-scapular lymph nodes (at 90%), bone marrow (at 85%), and ear pinnae (at 85%). This study demonstrated that the use of real-time PCR-HRM using the molecular marker hsp70 was a highly sensitive method for simultaneously detecting and identifying Leishmania species in different tissues taken from infected dogs. In addition, the usefulness of ear pinnae as easily accessible tissue for molecular diagnosis was emphasized.

Antitumor Immunity from Abdominal Flap-Embedded Therapeutic Cancer Vaccine.

BackgroundAbdominal flaps are routinely performed in clinic after primary mastectomy of breast cancer. However, cancer patients can still develop cancer recurrence and metastasis after surgery. In this study, we evaluated the feasibility of concurrent abdominal flap reconstruction and vaccine inoculation in the tissue for prevention and treatment of HER2-positive breast cancer.MethodsA murine model of metastatic HER2-positive breast cancer was generated by inoculating HER2-expressing TUBO tumor cells into both the mammary gland fat pad and left ventricle. Mammary gland fat pad with primary tumor was resected by mastectomy, and superficial inferior epigastric (SIE) vessel-based abdominal flap was performed for abdominal reconstruction. During the surgery, mice also received a single intra-flap treatment of a microparticulate-based cancer vaccine. Popliteal (Pop) and inguinal (Ing) lymph nodes (LN) were collected at different time points after vaccination, and activation of dendritic cells and T lymphocytes was evaluated with flow cytometry. ELISpot was also performed to measure HER2-specific T cells in splenocytes. In addition, infiltration of CD3+ T cells in brain metastatic nodules was analyzed with immunohistochemistry.ResultsFlow cytometry detected increased number of activated dendritic cells in lymph nodes in mice treated with cancer vaccine. ELISpot revealed abundant IFN-γ-expressing T cells in the spleen. Mice treated with abdominal flap-embedded cancer vaccine extended median survival by 9 days over the control group (p<0.05).ConclusionAbdominal flap-embedded cancer vaccine effectively stimulated systemic immune response and inhibited tumor progression in a murine model of HER2-positive breast cancer.

A case of malignant melanoma, a possible primary site in the digit, with systemic metastasis in a mini-Rex.

We report a case of systemic metastasis of malignant melanoma in a mini-Rex (Oryctolagus cuniculus). The animal presented with lameness of the right hind limb, swelling of popliteal lymph node, and a black mass on the first digit. Paralysis of hindlimbs and forelimbs, dysuria, and dysphagia progressed over time, and the rabbit died on day 35 from the first visit. At necropsy, many black lesions were observed in multiple organs including the marrow of most bones. Histopathologically, the tumor cells had highly atypical nuclei of various sizes and an abundant eosinophilic cytoplasm, and some cells contained melanin granules. These cells were positive for PNL2 and S-100, melanoma markers, by immunohistochemistry. This is the first report of malignant melanoma in a mini-Rex with severe malignancy and systemic metastasis including the bone marrow.

A case of a 4-year-old boy with difficult-to-diagnose skin infection with nontuberculous mycobacteria

Nontuberculous mycobacteria infections have become increasingly common in recent years and have even been confirmed in children. In addition to other organs, they can even affect the skin; nevertheless, in children lymphadenitis is the most common manifestation of the infection. The diagnosis of mycobacterial skin infection is based on patient history, clinical picture, histopathological changes, and tuberculin test result. Evidence of the causative agent in the lesion is confirmed with cultivation and PCR, two of the main tests that help determine the type of the causative mycobacteria. Here we report the case of a 4-year-old boy that presented with a few pink-to-livid papules and one plaque with a central crust on the skin of the left knee and an enlarged popliteal lymph node, highly suspicious of nontuberculous mycobacteria infection. Among the laboratory results, only a positive QuantiFERON and Mantoux test stood out. In addition, in the histopathological report, superficial and deep inflammatory elements were described, which could be due to an infection with nontuberculous mycobacteria. Despite negative cultivation and PCR, in agreement with a pediatric pulmonologist we decided to introduce antibiotic therapy for 6 months. Treatment was successful, we achieved regression of the skin lesions, and lymphadenitis was no longer present.

Comparative Study of Real-Time PCR (TaqMan Probe and Sybr Green), Serological Techniques (ELISA, IFA and DAT) and Clinical Signs Evaluation, for the Diagnosis of Canine Leishmaniasis in Experimentally Infected Dogs.

Canine leishmaniasis (CanL) diagnosis is not fully resolved. Currently, two specific methodologies are in continuous development, the detection of the parasite DNA or RNA in target organs and the detection of specific antibodies against Leishmania sp. For a correct diagnosis, it has been shown that the joint use of this type of test is necessary. In this work, a Sybr Green and a TaqMan Probe based on real time PCRs (qPCR) was performed for the detection of Leishmania sp. in order to correlate the results with clinicopathological and serological evaluations (IFA, ELISA and DAT) to propose an optimal biological sample to be used to detect the parasite in both early and late stages of the infection. A total of four samples were processed: conjunctival swabs, popliteal lymph node aspirates, bone marrow aspirates, and peripheral blood from experimentally infected dogs belonging to a larger study. Our results indicated that a single non-invasive sample (conjunctival swab) and the application of both types of qPCR would be reliable for determining Leishmania infection as well as the disease stage in dogs, thus avoiding bone marrow, lymph node aspirate or blood samples collection.

Comparison of normal hindlimb lymphatic systems in rats with detours present after lymphatic flow blockage.

In the present study, we aimed to identify the normal hindlimb lymphatic systems in rats and compare them with the detours after lymphatic flow blockage. The lymphatic systems of the hindlimbs of normal rats were investigated via lymphography using a near-infrared fluorescence imaging system. The lymphatic vessels were stained using Evans Blue. The lymphatic flow was blocked through lymphatic vessel ligation combined with inguinal and popliteal lymph node dissection. Detours that appeared after 30 days were visualized using lymphography and immunostaining with anti-podoplanin antibodies. Three main results were obtained in the present study. First, the deep medial system, the superficial medial system, a connection between the superficial and deep medial lymphatic systems, and the superficial lateral system, were elucidated. Second, three types of detours, namely the detour of the lateral abdomen, the detour to the lymphatic vessel near the midline of the abdomen, and the detour to the contralateral inguinal lymph node, were identified after lymphatic flow blockage. Lastly, detours were located in the fatty layer above the panniculus carnosus muscle and their lumina were wide. The histology suggested that the detour was a pre-collecting lymphatic vessel. Lymphatic routes in the rat hindlimbs after lymphatic flow blockage were different from those of the normal rat lymphatic system. It was suggested that the detour is a pre-collecting lymphatic vessel and that encouraging its development may be a new method of simple lymphatic drainage.

Comparison of shear-wave velocities obtained with shear-wave elastography of various peripheral lymph nodes in healthy Beagles

Abstract OBJECTIVE To compare shear-wave velocities (SWVs) with shear-wave elastography of various peripheral lymph nodes (LNs). ANIMALS 11 healthy Beagles. PROCEDURES For each dog, bilateral mandibular, medial retropharyngeal, superficial cervical, axillary, superficial inguinal, and popliteal LNs were evaluated with shear-wave elastography in sagittal and transverse scanning planes. Depth of each lymph node was recorded, and intra- and interobserver reliability was determined. RESULTS SWVs for all LNs were significantly higher in the sagittal scanning plane, compared with those in the transverse scanning plane. The SWV of the most superficial LN, the mandibular LN, was significantly higher, compared with that for the other LNs, except for the medial retropharyngeal LN. The SWV of the deepest LN, the medial retropharyngeal LN, was as high as that for the mandibular LN. Intra- and interobserver reliability was excellent. CONCLUSIONS AND CLINICAL RELEVANCE SWVs for normal peripheral LNs of Beagles may serve as a reference to compare with those for other breeds and diseased LNs. Scanning plane, LN depth, and interfering tissues between the LN and the transducer may affect SWV. Shear-wave elastography may not be operator dependent.

Comparison of shear-wave velocities obtained with shear-wave elastography of various peripheral lymph nodes in healthy Beagles.

To compare shear-wave velocities (SWVs) with shear-wave elastography of various peripheral lymph nodes (LNs). 11 healthy Beagles. For each dog, bilateral mandibular, medial retropharyngeal, superficial cervical, axillary, superficial inguinal, and popliteal LNs were evaluated with shear-wave elastography in sagittal and transverse scanning planes. Depth of each lymph node was recorded, and intra- and interobserver reliability was determined. SWVs for all LNs were significantly higher in the sagittal scanning plane, compared with those in the transverse scanning plane. The SWV of the most superficial LN, the mandibular LN, was significantly higher, compared with that for the other LNs, except for the medial retropharyngeal LN. The SWV of the deepest LN, the medial retropharyngeal LN, was as high as that for the mandibular LN. Intra- and interobserver reliability was excellent. SWVs for normal peripheral LNs of Beagles may serve as a reference to compare with those for other breeds and diseased LNs. Scanning plane, LN depth, and interfering tissues between the LN and the transducer may affect SWV. Shear-wave elastography may not be operator dependent.

Anatomy and nomenclature of tree shrew lymphoid tissues.

The immune response plays a key role in the disease development of the organism, while immune function serves as an important indicator for animal models evaluation. The tree shrew (Tupaia belangeri chinensis), as a new laboratory animal with a close genetic relationship with primates, has been used to construct various disease models. However, the immune system of tree shrews, especially anatomical descriptions of lymph nodes, is still relatively unknown. In this study, a total of 16 different lymph nodes were identified, including superficial lymph nodes and deep lymph nodes. Superficial lymph nodes were located in the head and neck region (submandibular lymph node, parotid lymph node, deep and superficial cervical lymph nodes) and at the forelimb (axillary and accessory axillary lymph nodes, subscapular lymph node) and hindlimb (popliteal, sciatic, and inguinal lymph nodes). Deep lymph nodes comprise mediastinal lymph nodes located in thoracic cavity and abdominal lymph nodes that are mainly located in each mesentery (mesenteric, gastric, pancreatic-duodenal, renal lymph nodes) or along the major vessels (iliac lymph nodes). In addition, we described the spleen and thymus of the tree shrew, as well as two lymphoid tissues in the top wall of the nasal cavity and the oropharynx. This study mainly describes the tree shrew immune system from an anatomical and histopathological perspective and provides fundamental research references for the establishment of various animal models of tree shrews.

Transcriptional inhibition by CDK7/9 inhibitor SNS-032 suppresses tumor growth and metastasis in esophageal squamous cell carcinoma

Metastasis is one of most lethal causes that confer a poor prognosis of patients with esophageal squamous cell carcinoma (ESCC), whereas there is no available target drug for metastatic ESCC currently. In this study, we aimed to determine whether the transcriptional inhibition by CDK7/9 inhibitor SNS-032 is activity against ESCC. MTT and soft agar assays were performed to examine the influence of SNS-032 on ESCC growth in vitro. Tumor xenograft in nude mice was used to assess the antitumor activity of SNS-032 in vivo. The roles of SNS-032 in ESCC metastasis were conducted by wound healing and transwell assays in vitro, and by a lung and a popliteal lymph node metastasis model in vivo. The results showed that CDK7 and CDK9 were highly expressed in ESCC cells; SNS-032 effectively inhibited cellular viability, abrogated anchorage-independent growth, and potentiated the sensitivity to cisplatin in ESCC cells in vitro and in vivo. In addition, SNS-032 induced a mitochondrial-dependent apoptosis of ESCC cells by reducing Mcl-1 transcription. SNS-032 also potently abrogated the abilities of ESCC cell migration and invasion through transcriptional downregulation of MMP-1. Importantly, SNS-032 remarkably inhibited the growth of ESCC xenograft, increased the overall survival, as well as diminished the lung and lymph node metastasis in nude mice. Taken together, our findings highlight that the CDK7/9 inhibitor SNS-032 is a promising therapeutic agent, and warrants a clinical trial for its efficacy in ESCC patients, even those with metastasis.

β-glucan from Aureobasidium pullulans augments the anti-tumor immune responses through activated tumor-associated dendritic cells

Dendritic cells (DCs) are recognized as the most potent antigen-presenting cells, capable of priming both naïve and memory T cells. Thus, tumor-resident DCs (tumor-associated DCs: TADCs) play a crucial role in the immune response against tumors. However, TADCs are also well known as a “double-edged sword” because an immunosuppressive environment, such as a tumor microenvironment, maintains the immature and tolerogenic properties of TADCs, resulting in the deterioration of the tumor. Therefore, it is essential to maintain and enhance the anti-tumoral activity of TADCs to aid tumor elimination. This study demonstrated the potential for tumor growth inhibition of Aureobasidium pullulan-derived β-glucan (AP-BG). Administration of AP-BG dramatically limited the development of different types of tumor cell lines transplanted into mice. Examination of the tumor-infiltrating leukocytes revealed that AP-BG caused high expression of co-stimulatory molecules on TADCs and enhanced the production of cytolytic granules as well as pro-inflammatory cytokines by the tumor-resident T cells. Furthermore, the syngeneic mixed lymphoid reaction assay and popliteal lymph node assay showed the significant ability of AP-BG to improve DCs’ antigen-specific priming of T cells in vitro and in vivo. Taken together, β-glucan might be an immune-potentiating adjuvant for cancer treatment. This highly widely-used reagent will initiate a new way to activate DC-targeted cancer immune therapy.

Autonomic Regulation of Nociceptive and Immunologic Changes in a Mouse Model of Complex Regional Pain Syndrome

Chronic pain frequently develops after limb injuries, and its pathogenesis is poorly understood. We explored the hypothesis that the autonomic nervous system regulates adaptive immune system activation and nociceptive sensitization in a mouse model of chronic post-traumatic pain with features of complex regional pain syndrome (CRPS). In studies sympathetic signaling was reduced using 6-hydroxydopamine (6-OHDA) or lofexidine, while parasympathetic signaling was augmented by nicotine administration. Hindpaw allodynia, unweighting, skin temperature, and edema were measured at 3 and 7 weeks after fracture. Hypertrophy of regional lymph nodes and IgM deposition in the skin of injured limbs were followed as indices of adaptive immune system activation. Passive transfer of serum from fracture mice to recipient B cell deficient (muMT) mice was used to assess the formation of pain-related autoantibodies. We observed that 6-OHDA or lofexidine reduced fracture-induced hindpaw nociceptive sensitization and unweighting. Nicotine had similar effects. These treatments also prevented IgM deposition, hypertrophy of popliteal lymph nodes, and the development of pronociceptive serum transfer effects. We conclude that inhibiting sympathetic or augmenting parasympathetic signaling inhibits pro-nociceptive immunological changes accompanying limb fracture. These translational results support the use of similar approaches in trials potentially alleviating persistent post-traumatic pain and, possibly, CRPS. PerspectiveSelective treatments aimed at autonomic nervous system modulation reduce fracture-related nociceptive and functional sequelae. The same treatment strategies limit pain-supporting immune system activation and the production of pro-nociceptive antibodies. Thus, the therapeutic regulation of autonomic activity after limb injury may reduce the incidence of chronic pain.

WIPI-1 inhibits metastasis and tumour growth via the WIPI-1-TRIM21 axis and MYC regulation in nasopharyngeal carcinoma

The metastatic rate of nasopharyngeal carcinoma (NPC) is the highest among head and neck tumours. Additionally, distant metastasis is the main cause of therapy failure and mortality in NPC. Thus, novel biomarkers are needed for designing new therapeutic strategies to improve the prognosis of this disease. In this study, qRT-PCR and western blotting revealed that the expression of the WD repeat domain phosphoinositide interacting 1 (WIPI-1) was markedly decreased in NPC cells and tissues. Furthermore, low WIPI-1 expression closely correlated with poor prognosis in NPC patients. In vitro functional experiments revealed that overexpression or knockdown of WIPI-1 repressed or facilitated the migration, colony formation, and proliferation of NPC cells. Consistent with the in vitro studies, WIPI-1 significantly inhibited tumour growth, invasion and metastasis in popliteal lymph node metastasis, lung metastasis, and xenograft mouse models in vivo. Mechanistically, WIPI-1 directly interacted with tripartite motif containing 21 (TRIM21) and enhanced starvation-induced autophagy by interacting with TRIM21 in NPC cells. Moreover, MYC gene expression was markedly increased in the WIPI-1 knockdown group, as demonstrated by RNA-seq analysis and qRT-PCR validation. Altogether, WIPI-1 acts as a tumour suppressor gene in NPC that inhibits tumour growth and metastasis. Targeting WIPI-1 may be a novel treatment approach for NPC.

Differential detection of metastatic and inflammatory lymph nodes using inflow-based vascular-space-occupancy (iVASO) MR imaging

PurposeTo investigate the potential value of inflow-based vascular-space-occupancy (iVASO) MR imaging in differentiating metastatic from inflammatory lymph nodes (LNs). MethodsTen female New Zealand rabbits with 2.5–3.0 kg body weight were studied. VX2 cells and egg yolk emulsion were inoculated into left and right thighs, respectively, to induce ten metastatic and ten inflammatory popliteal LNs. Conventional MRI and iVASO were performed 2 h prior to, and 10, 20 days after inoculation (D0, D10, D20). The short-axis diameter (S), short- to long-axis diameter ratio (SLR), and arteriolar blood volume (BVa) at each time point and their longitudinal changes of each model were recorded and compared. At D20, all rabbits were sacrificed to perform histological evaluation after the MR scan. ResultsThe mean values of S, SLR and BVa showed no significant difference between the two groups at D0 (P = 0.987, P = 0.778, P = 0.975). The BVa of the metastatic group was greater than that of the inflammatory at both D10 and D20 (P < 0.05; P < 0.001), whereas the S and SLR of the metastatic group were greater only at D20 (P < 0.001; P = 0.001). Longitudinal analyses showed that the BVa of the metastatic group increased at both D10 and D20 (P = 0.004; P = 0.001), while that of the inflammatory group only increased at D10 (P = 0.024). ConclusionThe BVa measured with iVASO has the potential to detect early metastatic LNs.

Creation of a rat lymphedema model using extensive lymph node dissection and circumferential soft tissue resection: Is this a reliable model?

The medical demand for lymphedema treatment is huge since the disease mechanism remains unclear, and management are difficult. Our purpose was to develop a reliable lymphedema model mimicking the clinical scenario and allows a microsurgical approach. Male Lewis rats weighing 400 to 450 g were used to create lymphedema with groin and popliteal lymph node dissection and creation of 5 mm circumferential skin defect (n=6). A skin incision was made and closed primarily for control group (n=5). Evaluation included indocyanine green (ICG) lymphangiography 1 and 2months postoperatively, volume difference between bilateral hindlimbs measured using micro-CT, and the skin was harvested for histological evaluation 2months postoperatively. Larger volume differences present in the lymphedema group (17.50 ± 7.76 vs. 3.73 ± 2.66%, p < .05). ICG lymphangiography indicated dermal backflow only in the lymphedema group. Increased thickness of the epidermis was noted in lymphedema group (28.50 ± 12.61 μm vs. 15.10 ± 5.41 μm, p < .0001). More CD45+ (35.6 ± 26.68 vs. 2.8 ± 4.23 cells/high power field [HPF], p < .0001), CD3+ (38.39 ± 20.17 vs. 9.73 ± 8.62 cells/HPF, p < .0001), and CD4+ cell infiltration (11.7 ± 7.71 vs. 2.0 ± 2.67 cells/HPF, p < .0001) were observed in the lymphedema group. Collagen type I deposition was more in the lymphedema group (0.15 ± 0.06 vs. 0.07 ± 0.03, p < .0005). A rat lymphedema model was successfully established. The model can be applied in lymphedema related research.

Shear wave velocity measurements obtained in different regions are repeatable for presumed normal canine lymph nodes: A pilot study.

Shear wave elastography (SWE) has been applied as a noninvasive method for predicting regional lymph node (LN) metastases in human and veterinary patients. However, published studies describing standardized protocols and repeatability of this technique are currently lacking. The objective of this prospective, pilot, observer agreement study was to determine whether different shear wave velocity (SWV) measurements obtained in different regions of presumed normal canine LNs would be repeatable. Two imagers consecutively performed shear wave elastography of submandibular, superficial inguinal, and popliteal LNs in 10, clinically healthy adult dogs. Ten elastograms of each LN were acquired by each imager. In each adequate elastogram, three regions of interest (ROIs) were placed in the softest and stiffest region of the LN. Additionally, one ROI was drawn covering the entire LN. In each ROI, mean, median, and maximum SWVs were calculated. Mean values for the mean, median, and maximum SWVs varied from 2.33 to 3.10 m/s, 2.32 to 3.10 m/s, and 2.61 to 4.09 m/s, respectively. Intra- and interobserver agreements were acceptable. Superficial inguinal LNs demonstrated the highest intra- and interobserver agreement, followed by the popliteal and the submandibular LNs, respectively. Using the different measurements (mean, median, or maximum SWVs) had no significant effect on the intra- and interobserver variability, neither did the region (softest, stiffest, or entire LN). Findings indicated that all evaluated measurements and regions could be used to obtain reliable elastography data of presumed normal canine LNs. Clinical trials in dogs with cancer are necessary to compare SWVs of metastatic LNs with the reported SWVs and evaluate whether various measurements and regions can also be used in metastatic LNs.

Ex vivo Demonstration of Functional Deficiencies in Popliteal Lymphatic Vessels From TNF-Transgenic Mice With Inflammatory Arthritis.

Background: Recent studies demonstrated lymphangiogenesis and expansion of draining lymph nodes during chronic inflammatory arthritis, and lymphatic dysfunction associated with collapse of draining lymph nodes in rheumatoid arthritis (RA) patients and TNF-transgenic (TNF-Tg) mice experiencing arthritic flare. As the intrinsic differences between lymphatic vessels afferent to healthy, expanding, and collapsed draining lymph nodes are unknown, we characterized the ex vivo behavior of popliteal lymphatic vessels (PLVs) from WT and TNF-Tg mice. We also interrogated the mechanisms of lymphatic dysfunction through inhibition of nitric oxide synthase (NOS).Methods: Popliteal lymph nodes (PLNs) in TNF-Tg mice were phenotyped as Expanding or Collapsed by in vivo ultrasound and age-matched to WT littermate controls. The PLVs were harvested and cannulated for ex vivo functional analysis over a relatively wide range of hydrostatic pressures (0.5–10 cmH2O) to quantify the end diastolic diameter (EDD), tone, amplitude (AMP), ejection fraction (EF), contraction frequency (FREQ), and fractional pump flow (FPF) with or without NOS inhibitors Data were analyzed using repeated measures two-way ANOVA with Bonferroni’s post hoc test.Results: Real time videos of the cannulated PLVs demonstrated the predicted phenotypes of robust vs. weak contractions of the WT vs. TNF-Tg PLV, respectively. Quantitative analyses confirmed that TNF-Tg PLVs had significantly decreased AMP, EF, and FPF vs. WT (p < 0.05). EF and FPF were recovered by NOS inhibition, while the reduction in AMP was NOS independent. No differences in EDD, tone, or FREQ were observed between WT and TNF-Tg PLVs, nor between Expanding vs. Collapsed PLVs.Conclusion: These findings support the concept that chronic inflammatory arthritis leads to NOS dependent and independent draining lymphatic vessel dysfunction that exacerbates disease, and may trigger arthritic flare due to decreased egress of inflammatory cells and soluble factors from affected joints.

Decoy Immunization for Mice, Rats, and Hamsters.

Decoy immunization relies on misdirecting the T and B cells of the immune system away from immunogenic regions against which one does not want to generate antibodies, allowing the desired region to be profiled. Essentially, it involves immunizing animals with two forms of the targeted protein on opposite sides of the body. One form, with the region of interest removed (i.e., a modified protein), is injected into the left side of animal, whereas the intact protein of interest is injected into the right side. In theory, undesired B cells are drawn to the left side of the animal, leaving the desired B cells to be drawn to the right (specifically, in the procedure presented here, to the right popliteal lymph node).

Inhibition of glycolysis in the presence of antigen generates suppressive antigen-specific responses and restrains rheumatoid arthritis in mice

Dendritic cells (DCs) rely on glycolysis for their energy needs to induce pro-inflammatory antigen-specific immune responses. Therefore, inhibiting DC glycolysis, while presenting the self-antigen, may prevent pro-inflammatory antigen-specific immune responses. Previously we demonstrated that microparticles with alpha-ketoglutarate (aKG) in the polymer backbone (paKG MPs) were able to generate anti-inflammatory DCs by sustained delivery of the aKG metabolite, and by modulating energy metabolism of DCs. Herein, we demonstrate that paKG MPs-based delivery of a glycolytic inhibitor, PFK15, using paKG MPs induces anti-inflammatory DCs (CD86LoMHCII+) by down-regulating glycolysis, CD86, tnf and IL-6 genes, while upregulating oxidative phosphorylation (OXPHOS) and mitochondrial genes. Furthermore, paKG MPs delivering PFK15 and a self-antigen, collagen type II (bc2), in vivo, in a collagen-induced autoimmune arthritis (CIA) mouse model, normalized paw inflammation and arthritis score, by generating antigen-specific immune responses. Specifically, these formulations were able to reduce activation of DCs in draining lymph nodes and impressively generated proliferating bc2-specific anti-inflammatory regulatory T cells in joint-associated popliteal lymph nodes. These data strongly suggest that sustained glycolytic inhibition of DCs in the presence of an antigen can induce antigen-specific immunosuppressive responses, therefore, generating a technology that can be applicable for treating autoimmune diseases.

Core-Needle Biopsy Efficacy in Histopathology Diagnosis of Canine Multicentric Lymphoma

Core-needle biopsy (CNB) is now widely used to diagnose and classify human lymphoma. Few studies, however, have reported the use of CNB in veterinary medicine for dogs with suspected lymphoma. The present study evaluated the efficacy of CNB sample collection to diagnose canine multicentric lymphoma morphology. 16 dogs of varying breeds with generalized peripheral lymphadenopathy and cytopathologic features suggestive of lymphoma were enrolled in the study. CNB followed by lymphadenectomy of the left popliteal lymph node was performed. Both samples underwent histopathologic evaluation. Among the 16 dogs, 14 had lymphoma. CNB was positive for lymphoma in 12 (85.7%) of these 14 dogs. CNB analysis for diagnosing dogs with multicentric lymphoma exhibited a sensitivity of 86% and a specificity of 100%. CNB was effective in generalized lymphadenopathies toward achieving a differential diagnosis and microscopic evaluation of multicentric lymphoma in dogs, revealing essential features for morphologic classification, such as cell size, histopathologic type, and grade.