Automated detection and delineation of lymph nodes in haematoxylin & eosin stained digitised slides.

Treatment of patients with oesophageal and gastric cancer (OeGC) is guided by disease stage, patient performance status and preferences. Lymph node (LN) status is one of the strongest prognostic factors for OeGC patients. However, survival varies between patients with the same disease stage and LN status. We recently showed that LN size from patients with OeGC might also have prognostic value, thus making delineations of LNs essential for size estimation and the extraction of other imaging biomarkers. We hypothesized that a machine learning workflow is able to: (1) find digital H&E stained slides containing LNs, (2) create a scoring system providing degrees of certainty for the results, and (3) delineate LNs in those images. To train and validate the pipeline, we used 1695 H&E slides from the OE02 trial. The dataset was divided into training (80%) and validation (20%). The model was tested on an external dataset of 826 H&E slides from the OE05 trial. U-Net architecture was used to generate prediction maps from which predefined features were extracted. These features were subsequently used to train an XGBoost model to determine if a region truly contained a LN. With our innovative method, the balanced accuracies of the LN detection were 0.93 on the validation dataset (0.83 on the test dataset) compared to 0.81 (0.81) on the validation (test) datasets when using the standard method of thresholding U-Net predictions to arrive at a binary mask. Our method allowed for the creation of an "uncertain" category, and partly limited false-positive predictions on the external dataset. The mean Dice score was 0.73 (0.60) per-image and 0.66 (0.48) per-LN for the validation (test) datasets. Our pipeline detects images with LNs more accurately than conventional methods, and high-throughput delineation of LNs can facilitate future LN content analyses of large datasets.

Merkel cell carcinoma: Epidemiology, disease presentation, and current clinical practice outcomes.

Using the National Cancer Database, we introduce the findings of a retrospective investigation of the largest cohort of cases with Merkel cell carcinoma (N = 20,829). A decreasing proportion of stage I (P = .0004) and stage II (P = .0065) Merkel cell carcinoma among skin cancers was complemented by an increasing proportion of stage III disease (P < .0001). A predominance of non-Hispanic White (96.4%), male (62.6%) patients with a mean age of 74.5 ± 10.8 years and Medicare coverage (73.5%) was observed. Stage I was the most common presenting stage at diagnosis (29.2%), followed by stages II (12.7%), III (11.0%), and IV (3.8%). Most Merkel cell carcinoma tumors grew outside the head and neck (53.4%) and showed a nodular growth pattern (66.0%) but no extracapsular lymph node (90.5%) or lymphovascular involvement (63.8%). Narrow-margin excision and radiation therapy (RT) were used in 75.2% and 56.3% of tumors, respectively. Wide-margin excision lead to improved overall survival (P < .001) versus narrow-margin excision, particularly in stage III (difference in the median overall survival rate [ΔmOS], 23.7 months; P < .001). RT showed a significant OS benefit (P =.006), most pronounced in stage II (ΔmOS, 37.8 months) followed by stage I (ΔmOS, 16.1 months; P < .001). The survival benefit with primary-site RT (ΔmOS, 24.0 months) was higher than that with primary-site/lymph node RT (ΔmOS, 5.2 months; P < .001). Wide-margin excision independently predicted improved OS (hazard ratio, 0.577; 95% CI, 0.403-0.826; P = .003) versus narrow-margin excision and RT predicted better OS (hazard ratio, 0.608; 95% CI, 0.424-0.873; P = .007) versus no RT on multivariable analysis.

A Phenotypic Analysis of Involucrin-Membrane-Bound Ovalbumin Mice after Adoptive Transfer of Ovalbumin-Specific CD8+ T Cells.

To investigate the mechanism of autoimmunity and peripheral tolerance in the skin, several transgenic mouse strains expressing membrane-bound ovalbumin (mOVA) as an epidermal self-antigen under the control of keratinocyte-specific promotors, such as keratin 5 and keratin 14, were employed in combination with adoptive transfer of CD8+ T cells from OT-I mice (OT-I T cells) that recognize an ovalbumin-derived peptide. However, these strains showed bodyweight loss and required additional inflammatory stimuli, such as γ-irradiation and tape-stripping, to induce skin inflammation. In this study, we generated a mouse strain expressing mOVA under the control of human involucrin promoter (involucrin-mOVA mice). In contrast to previous strains, involucrin-mOVA mice spontaneously developed skin inflammation after the transfer of OT-I T cells in the absence of external stimuli without significant bodyweight loss. We focused on the skin infiltration process of OT-I T cells and found that transferred OT-I T cells accumulated around the hair follicles in the early phase of skin inflammation, and in the later phase, the skin inflammation spontaneously resolved despite the remaining OT-I T cells in the skin. Our involucrin-mOVA mice will provide a promising tool to investigate the pathogenesis and the tolerance mechanisms of cytotoxic skin autoimmunity.

Primary pulmonary synovial sarcoma.

Objective: Primary pulmonary synovial sarcoma (PPSS) is extremely rare. This study aims to identify the clinicopathologic and therapeutic factors determining survival in PPSS. Methods: We performed a retrospective analysis of 121 patients from the Surveillance, Epidemiology, and End Results Database as well as 12 patients from our own institution diagnosed with PPSS. Patient survival was evaluated using the Kaplan-Meier method. Results: The median survival time for 12 PPSS patients in our institution was 78 months. Postoperative chemotherapy (P = .027 for overall survival and P = .035 for disease-specific survival) was associated with superior survival, whereas pneumonectomy (P = .011 for overall survival and P = .006 for disease-specific survival) was associated with worse survival. Single lobe involvement (P = .022) and the absence of lymph node involvement (P = .045) were associated with improved disease-specific survival and overall survival, respectively. In the Surveillance, Epidemiology, and End Results Database, the median survival time was 23 months. Significantly superior survival was observed in patients with earlier American Joint Committee on Cancer stage (Ⅰ-Ⅱ) (P < .001 for both overall survival and disease-specific survival). Patients who were diagnosed within the recent decade did not achieve a better survival (P = .599 for overall survival and P = .596 for disease-specific survival). Conclusions: PPSS was aggressive with a very poor prognosis. The seventh American Joint Committee on Cancer stage might aid in predicting survival. Pneumonectomy and lymph node involvement might be associated with worse survival, whereas single lobe involvement and postoperative chemotherapy might be associated with improved survival.

Interlobar division using vessel-sealing system in robot-assisted pulmonary lobectomy.

Objective: We investigated the safety of a novel interlobar fissure division technique using the da Vinci vessel sealing system in robot-assisted pulmonary lobectomy. Methods: The medical records of patients who underwent robotic pulmonary lobectomy with node dissection for primary lung cancer between 2018 and 2020 were reviewed. The inclusion criteria were fulfilled by 111 patients, whose perioperative factors and postoperative results were compared with those previously reported. Furthermore, the new robotic lung interlobar division technique using the da Vinci vessel sealing system without a robotic stapler was evaluated in patients with low-grade incomplete fissure. We considered the Craig and Walker classification of lung fissures grades 1 and 2 as a good adaptation for the vessel sealing system interlobar fissure division. Results: The vessel sealing system group had shorter mean operative and console times (P = .03 and P = .01, respectively) and lesser median intraoperative blood loss (20 mL vs 50 mL; P = .01). The vessel sealing system group had lower surgical complication rates (2.2% vs 20.0%; P = .01). The incidence of persistent postoperative air leak was lower (0% vs 10.0%; P = .06), and fewer robotic stapler cartridges were used during surgery (3.4 vs 5.6; P < .001) in the vessel sealing system group than in the stapler group. Conclusions: We report the safety of using the da Vinci vessel sealing system as an alternative to the use of robotic staples for interlobar fissure division in robot-assisted pulmonary lobectomy. This technique seems easy and feasible though limited to the low-grade incomplete fissure.

Integrative lymph node-mimicking models created with biomaterials and computational tools to study the immune system.

The lymph node (LN) is a vital organ of the lymphatic and immune system that enables timely detection, response, and clearance of harmful substances from the body. Each LN comprises of distinct substructures, which host a plethora of immune cell types working in tandem to coordinate complex innate and adaptive immune responses. An improved understanding of LN biology could facilitate treatment in LN-associated pathologies and immunotherapeutic interventions, yet at present, animal models, which often have poor physiological relevance, are the most popular experimental platforms. Emerging biomaterial engineering offers powerful alternatives, with the potential to circumvent limitations of animal models, for in-depth characterization and engineering of the lymphatic and adaptive immune system. In addition, mathematical and computational approaches, particularly in the current age of big data research, are reliable tools to verify and complement biomaterial works. In this review, we first discuss the importance of lymph node in immunity protection followed by recent advances using biomaterials to create in vitro/vivo LN-mimicking models to recreate the lymphoid tissue microstructure and microenvironment, as well as to describe the related immuno-functionality for biological investigation. We also explore the great potential of mathematical and computational models to serve as in silico supports. Furthermore, we suggest how both in vitro/vivo and in silico approaches can be integrated to strengthen basic patho-biological research, translational drug screening and clinical personalized therapies. We hope that this review will promote synergistic collaborations to accelerate progress of LN-mimicking systems to enhance understanding of immuno-complexity.

In Vivo Longitudinal Tracking of Lymphangiogenesis and Angiogenesis in Cutaneous Melanoma Mouse Model Using Multifunctional Optical Coherence Tomography.

Melanoma is a high-risk skin cancer because it tends to metastasize early and ultimately leads to death. In this study, we introduced a noninvasive multifunctional optical coherence tomography (MFOCT) for the early detection of premetastatic pathogenesis in cutaneous melanoma by label-free imaging of microstructures (i.e., providing the thickness and the scattering information) and microcirculation (i.e., providing depth-resolved angiography and lymphangiography). Using MFOCT-based approaches, we presented an in vivo longitudinal observation of the tumor microenvironment in Braf V600E/V600E ;Pten -/- mice with inducible melanoma monitored for 42 days. Quantitative analysis of MFOCT images identified an increased number of lymphatic and vascular vessels during tumor progression and faster lymphangiogenesis (beginning on day 21) than angiogenesis (beginning on day 28) in the melanoma microenvironment. We further observed lymphatic vessel enlargement from the first week of melanoma development, implying tumor cells interacting with the vessels and increased likelihood of metastasis. MFOCT identified cutaneous melanoma‒associated angiogenesis and lymphangiogenesis before the possible visual perception of the tumor (≥42 days) and before metastasis could be diagnosed using micropositron emission tomography (35 days). Thus, the proposed quantitative analysis using MFOCT has the potential for early detection of cutaneous melanoma progression or prediction of metastatic melanoma in a mouse model. However, retrospective and extensive experiments still need to be performed in the future to confirm the value of MFOCT in clinical application.

TNF-α and IL-1ß Do Not Induce Langerhans Cell Migration by Inhibiting TGFß Activation.

In the skin, Langerhans cells (LCs) require autocrine latent TGFß that is transactivated by the integrins ανß6 and ανß8 expressed by keratinocytes (KCs) for long-term epidermal retention. Selective expression of a ligand-independent, constitutively active form of TGFßR1 inhibits LC migration during homeostasis and in response to UVB exposure. In this study, we found that LC migration in response to inflammatory stimuli was also inhibited by ligand-independent TGFßR1 signaling. Contrary to UVB stimulation, which reduced KC expression of ανß6, in vitro and in vivo exposure to TNF-α or IL-1ß increased ανß6 transcript and protein expression by KCs. This resulted in increased KC-mediated transactivation of latent TGFß. Expression of ανß8 was largely unchanged. These findings show that ligand-independent TGFßR1 signaling in LCs can overcome inflammatory migration stimuli, but reduced KC-mediated transactivation of latent TGFß by KCs may only drive LC migration during homeostasis and in response to UV stimulation.

A quantitative systems pharmacology model for acute viral hepatitis B.

Hepatitis B liver infection is caused by hepatitis B virus (HBV) and represents a major global disease problem when it becomes chronic, as is the case for 80-90% of vertical or early life infections. However, in the vast majority (>95%) of adult exposures, the infected individuals are capable of mounting an effective immune response leading to infection resolution. A good understanding of HBV dynamics and the interaction between the virus and immune system during acute infection represents an essential step to characterize and understand the key biological processes involved in disease resolution, which may help to identify potential interventions to prevent chronic hepatitis B. In this work, a quantitative systems pharmacology model for acute hepatitis B characterizing viral dynamics and the main components of the innate, adaptive, and tolerant immune response has been successfully developed. To do so, information from multiple sources and across different organization levels has been integrated in a common mechanistic framework. The final model adequately describes the chronology and plausibility of an HBV-triggered immune response, as well as clinical data from acute patients reported in the literature. Given the holistic nature of the framework, the model can be used to illustrate the relevance of the different immune pathways and biological processes to ultimate response, observing the negligible contribution of the innate response and the key contribution of the cellular response on viral clearance. More specifically, moderate reductions of the proliferation of activated cytotoxic CD8+ lymphocytes or increased immunoregulatory effects can drive the system towards chronicity.

Repeated treatment with gefitinib and alectinib in a patient with multiple EGFR-mutant and ALK-mutant lung adenocarcinomas: A case report.

Multiple EGFR-mutant and ALK-mutant lung cancers are rare, and standard treatment has not been established because of the small number of cases. A 79-year-old man was found to harbor nodular shadows in right S1, right S5, and left S3. He was surgically diagnosed with stage IIB (pT3N0M0) EGFR G719X-mutant lung adenocarcinoma in left S3 and stage IA1 (pT1aN0M0) ALK-mutant lung adenocarcinoma in right S5. Owing to the relapse of the EGFR-mutant adenocarcinoma, gefitinib treatment was commenced 3 months postoperatively. The tumor shrank temporarily; however, the nodular shadow in the right S1 and #3a lymph nodes were found to increase in size. He was diagnosed with adenosquamous carcinoma in right S1 and relapsing ALK-mutant adenocarcinoma in #3a lymph node. Gefitinib treatment was continued, but due to a renewed increase in the size of the #3a lymph node, the drug was changed to alectinib 16 months postoperatively. Subsequently, the EGFR-mutant adenocarcinomas were found to increase in left S1 despite the decrease in the #3a lymph node size. Nineteen months after the first surgery, the treatment was changed to gefitinib, and repeated treatment with this drug and alectinib administered every 2 months was continued. This approach enabled 39 months of progression-free survival, and no serious adverse events were observed.

Intensity modulated radiotherapy for anal canal squamous cell carcinoma: A 16-year single institution experience.

INTRODUCTION: To report long-term efficacy and adverse events (AEs) associated with intensity modulated radiotherapy (IMRT) for patients with anal canal squamous cell carcinoma (ASCC). MATERIALS AND METHODS: This was a retrospective review of patients with ASCC who received curative-intent IMRT and concurrent chemotherapy (98%) between 2003 and 2019. Overall survival (OS), colostomy-free survival (CFS), and progression-free survival (PFS) were estimated using the Kaplan-Meier method. The cumulative incidence of local recurrence (LR), locoregional recurrence (LRR), and distant metastasis (DM) were reported. Acute and late AEs were recorded per National Cancer Institute Common Terminology Criteria for AEs. RESULTS: 127 patients were included. The median patient age was 63 years (interquartile range [IQR] 55-69) and 79% of patients were female. 33% of patients had T3-4 disease and 68% had clinically involved pelvic or inguinal lymph nodes (LNs).The median patient follow-up was 47 months (IQR: 28-89 months). The estimated 4-year OS, CFS, and PFS were 81% (95% confidence interval [CI]: 73%-89%), 77% (95% CI: 68%-86%), and 78% (95% CI: 70%-86%), respectively. The 4-year cumulative incidences of LR, LRR, and DM were 3% (95% CI: 1%-9%), 9% (95% CI: 5%-17%), and 10% (95% CI: 6%-18%), respectively. Overall treatment duration greater than 39 days was associated with an increased risk of LRR (Hazard Ratio [HR]: 5.2, 95% CI: 1.4-19.5, p = 0.015). The most common grade 3+ acute AEs included hematologic (31%), gastrointestinal (GI) (17%), dermatologic (16%), and pain (15%). Grade 3+ late AEs included: GI (3%), genitourinary (GU) (2%), and pain (1%). Current smokers were more likely to experience grade 3+ acute dermatologic toxicity compared to former or never smokers (34% vs. 7%, p < 0.001). CONCLUSIONS: IMRT was associated with favorable toxicity rates and long-term efficacy. These data support the continued utilization of IMRT as the preferred treatment technique for patients with ASCC.

Residual lymph node disease and mortality following neoadjuvant chemoradiation and curative esophagectomy for distal esophageal adenocarcinoma.

Objectives: Neoadjuvant chemoradiation has been shown to improve survival in locally advanced esophageal and gastroesophageal junction cancer. The purpose of our study was to examine the effects of posttreatment persistent lymph node (LN) disease on overall survival (OS) and recurrence in patients with esophageal adenocarcinoma after neoadjuvant chemoradiation as well as the effect of LN harvest and the potential benefit of adjuvant chemotherapy. Methods: The records of patients who underwent esophagectomy in our hospital from January 2005 until December 2016 were analyzed. Our study group consisted of 509 patients. Results: Patient groups were created based on pathologic staging after esophagectomy (ypT N) as 22.0% of patients were ypT0 N0, 46.2% had incomplete response only at the primary tumor level (ypT + N0), and 31.8% had at least 1 metastatic lymph node (ypTx N+). Median OS was 58.3 months. The ypTx N+ group was divided into ypTx N1 and ypTx N2 or N3 subgroups based on the number of metastatic lymph nodes. The OS between the 2 groups was not significantly different (median OS, 37.6 vs 29.8 months; P = .097). The disease-free survival did show a statistically significant difference (median disease-free survival, 27.6 vs 13.7 months; P = .007). The LN harvest was not found to be significantly associated with OS. However, administration of adjuvant chemotherapy was a significant prognosticator for increased OS (hazard ratio, 0.590; P = .043). Conclusions: Our results demonstrate that residual LN disease after neoadjuvant chemoradiation is associated with increased mortality. Adjuvant chemotherapy, but not number of LNs resected, was correlated with increased OS in this subset of patients.

Fat fractions from high-resolution 3D radial Dixon MRI for predicting metastatic axillary lymph nodes in breast cancer patients.

PURPOSE: To assess diagnostic performance of fat fractions (FF) from high-resolution 3D radial Dixon MRI for differentiating metastatic and non-metastatic axillary lymph nodes in breast cancer patients. METHOD: High-resolution 3D radial Dixon MRI was prospectively performed on 1.5 T in 70 biopsy-verified breast cancer patients. 35 patients were available for analysis with histopathologic and imaging data. FF images were calculated as fat / in-phase. Two radiologists measured lymph node FF and assessed morphological features in one ipsilateral and one contralateral lymph node in consensus. Diagnostic performance of lymph node FF and morphological criteria were compared using histopathology as reference. RESULTS: 22 patients had metastatic axillary lymph nodes. Mean lymph node FF were 0.20 ±â€¯0.073, 0.31 ±â€¯0.079, and 0.34 ±â€¯0.15 (metastatic, non-metastatic ipsi- and non-metastatic contralateral lymph nodes, respectively). Metastatic lymph node FF were significantly lower than non-metastatic ipsi- (p <  0.001) and contralateral lymph nodes (p <  0.001). Area under the receiver operating characteristics curve for lymph node FF was 0.80 compared to 0.76 for morphological criteria (p =  0.29). Lymph node FF yielded sensitivity 0.91, specificity 0.69, positive predictive value (PPV) 0.83, and negative predictive value (NPV) 0.82, while morphological criteria yielded sensitivity 0.91, specificity 0.62, PPV 0.80, and NPV 0.80 (p =  0.71). Combining lymph node FF and morphological criteria increased diagnostic performance with sensitivity 1.00, specificity 0.67, PPV 0.86, NPV 1.00, and AUC 0.83. CONCLUSIONS: Lymph node FF from high-resolution 3D Dixon images are a promising quantitative indicator of metastases in axillary lymph nodes.

Hyperbilirubinemia predicts the infectious complications after esophagectomy for esophageal cancer.

BACKGROUND: Surgical stress and inflammation can cause hyperbilirubinemia, which sometimes occurs after esophagectomy for esophageal cancer (EC). The aim of this study was to elucidate the clinical significance of postoperative hyperbilirubinemia in the management of EC patients. MATERIALS AND METHODS: We retrospectively reviewed records of 81 EC patients who underwent esophagectomy from 2009 to 2014. We compared the clinicopathological and perioperative factors, including the presence of hyperbilirubinemia (total bilirubin ≥1.5 mg/dL), between patients with postoperative infectious complications (PIC group) and those without (Non-PIC group). RESULTS: PIC developed in 52 patients (64.2%). There were significant differences in incidence of postoperative hyperbilirubinemia between the PIC group and the non-PIC group (34.6% vs. 3.4%, P = 0.002), as well as the approach of esophagectomy (P = 0.045), the surgical duration (469 vs. 389 min, P < 0.001), the amount of blood loss (420 vs. 300 mL, P = 0.018), the frequency of intraoperative blood transfusions (32.7% vs. 6.9%, P = 0.012) and the peak postoperative C-reactive protein level (17.3 vs. 8.6 mg/dL, P = 0.007). Multivariate analysis revealed hyperbilirubinemia was independently associated with the occurrence of PICs (odds ratio: 38.6, P = 0.010). The median time to the diagnosis of hyperbilirubinemia was significantly shorter than that of PICs (3.0 vs. 4.5 days, P = 0.025). CONCLUSIONS: Postoperative hyperbilirubinemia was associated with the occurrence of PICs and frequently occurred before any PICs become apparent. More attention should be paid to the serum bilirubin level in the management after esophagectomy for EC.

CCR2+ Monocyte-Derived Infiltrating Macrophages Are Required for Adverse Cardiac Remodeling During Pressure Overload.

Although chronic inflammation is a central feature of heart failure (HF), the immune cell profiles differ with different underlying causes. This suggests that for immunomodulatory therapy in HF to be successful, it needs to be tailored to the specific etiology. Here, the authors demonstrate that monocyte-derived C-C chemokine receptor 2 (CCR2)+ macrophages infiltrate the heart early during pressure overload in mice, and that blocking this response either pharmacologically or with antibody-mediated CCR2+ monocyte depletion alleviates late pathological left ventricular remodeling and dysfunction, T-cell expansion, and cardiac fibrosis. Hence, suppression of CCR2+ monocytes/macrophages may be an important immunomodulatory therapeutic target to ameliorate pressure-overload HF.

A protocol for generating induced T cells by reprogramming B cells in vivo.

Obtaining T cells by reprogramming is one of the major goals in regenerative medicine. Here, we describe a protocol for generating functional T cells from Hoxb5-expressing pro/pre-B cells in vivo. This protocol includes the construction of Hoxb5 recombinant plasmids, retroviral packaging, isolation and viral transduction of pro/pre-B cells, cell transplantation, and phenotypic analysis of induced T cells. The procedure is reproducible and straightforward, providing an approach for generating induced T cells for translational research.

Squamous cell carcinoma of the urinary bladder: Systematic review of clinical characteristics and therapeutic approaches.

OBJECTIVE: To highlight the current understanding of the epidemiology, clinicopathological characteristics, and management of squamous cell carcinoma (SCC) of the bladder, as it accounts for 2-5% of bladder tumours, with a focus on non-bilharzial-associated SCC (NB-SCC). The standard treatment for bladder SCC remains radical cystectomy (RC). We present an updated clinical profile of bladder SCC and a review of NB-SCC therapeutic approaches, including RC, neoadjuvant and adjuvant treatments, radiotherapy, chemotherapy, and immunotherapy. METHODS: Using search terms relating to SCC, urinary bladder, and treatment modalities, we performed a search of the PubMed and Embase databases to identify NB-SCC treatment approaches and outcomes. Peer-reviewed English language reports from 1975 to present assessing SCC management were included. Two authors independently screened and extracted the data. RESULTS: Of the 806 articles screened, 10 met the pre-defined inclusion criteria. RC was performed in seven of the 10 studies. Although radiotherapy alone yielded poor outcomes, preoperative radiotherapy and RC were associated with improved survival. There is little evidence supporting the use of chemotherapy in NB-SCC, and its efficacy in relation to RC is not known. CONCLUSION: Based on current literature, there is insufficient evidence to provide a treatment recommendation for NB-SCC. Whilst RC is the standard of care, the role of preoperative radiotherapy should be revisited and compared to RC alone. Additional studies incorporating multimodal approaches, contemporary radiation techniques, and systemic therapies are warranted. Immunotherapy as a treatment for bladder SCC has yet to be investigated.

Adjuvant-enhanced CD4 T Cell Responses are Critical to Durable Vaccine Immunity.

Protein-based vaccines offer a safer alternative to live-attenuated or inactivated vaccines but have limited immunogenicity. The identification of adjuvants that augment immunogenicity, specifically in a manner that is durable and antigen-specific, is therefore critical for advanced development. In this study, we use the filovirus virus-like particle (VLP) as a model protein-based vaccine in order to evaluate the impact of four candidate vaccine adjuvants on enhancing long term protection from Ebola virus challenge. Adjuvants tested include poly-ICLC (Hiltonol), MPLA, CpG 2395, and alhydrogel. We compared and contrasted antibody responses, neutralizing antibody responses, effector T cell responses, and T follicular helper (Tfh) cell frequencies with each adjuvant's impact on durable protection. We demonstrate that in this system, the most effective adjuvant elicits a Th1-skewed antibody response and strong CD4 T cell responses, including an increase in Tfh frequency. Using immune-deficient animals and adoptive transfer of serum and cells from vaccinated animals into naïve animals, we further demonstrate that serum and CD4 T cells play a critical role in conferring protection within effective vaccination regimens. These studies inform on the requirements of long term immune protection, which can potentially be used to guide screening of clinical-grade adjuvants for vaccine clinical development.