Detection of SARS-CoV-2 RNA by In Situ Hybridization in Lung-Cancer Cells Metastatic to Brain and in Adjacent Brain Parenchyma.

The mechanisms by which severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) may spread to the human brain are poorly understood, and the infection of cancer cells in the brain by SARS-CoV-2 in Coronavirus disease 2019 (COVID-19) patients has been the subject of only one previous case report. Here, we report the detection of SARS-CoV-2 RNA by in situ hybridization in lung-cancer cells metastatic to the brain and adjacent brain parenchyma in a 63-year-old male patient with COVID-19. These findings suggest that metastatic tumors may transport the virus from other parts of the body to the brain or may break down the blood-brain barrier to allow for the virus to spread to the brain. These findings confirm and extend previous observations that cancer cells in the brain can become infected by SARS-CoV-2 in patients with COVID-19 and raise the possibility that SARS-CoV-2 can have a direct effect on cancer growth and outcome.

Host Molecules That Promote Pathophysiology of Ocular Herpes.

Herpes simplex virus type-1 (HSV-1) is a human virus that causes lifelong infections in a large population worldwide. Recurrence of HSV-1 from latency in trigeminal ganglion (TG) is the trigger of the morbidities seen with this virus. In addition to causing fever blisters and cold sores, occasionally the virus can also cause corneal lesions resulting in blindness in untreated individuals. Several host cell proteins play important roles in HSV-1 infection of the eye. HSV-1 enters into the corneal epithelial cells via its interactions with cell surface receptors. In parallel, the Toll-like receptors sense viral invasion and activate defense mechanisms to fight the infection. New data shows that Optineurin, a host autophagy receptor is also activated to degrade viral particles. In contrast, activation of heparanase, a host enzyme, induces an immune-inflammatory response, which triggers pro-inflammatory and pro-angiogenic environment and ultimately results in many of the clinical features seen with HSV-1 infection of the cornea. Rarely, HSV-1 can also spread to the central nervous system causing serious diseases. In this review, we summarize the latest knowledge on host molecules that promote pathophysiological aspects of ocular herpes.

OPTN is a host intrinsic restriction factor against neuroinvasive HSV-1 infection.

Fast-replicating neurotropic herpesviruses exemplified by herpes simplex virus-1 (HSV-1) naturally infect the central nervous system (CNS). However, most individuals intrinsically suppress the virus during a primary infection and preclude it from significantly damaging the CNS. Optineurin (OPTN) is a conserved autophagy receptor with little understanding of its role in neurotropic viral infections. We show that OPTN selectively targets HSV-1 tegument protein, VP16, and the fusion glycoprotein, gB, to degradation by autophagy. OPTN-deficient mice challenged with HSV-1 show significant cognitive decline and susceptibility to lethal CNS infection. OPTN deficiency unveils severe consequences for recruitment of adaptive immunity and suppression of neuronal necroptosis. Ocular HSV-1 infection is lethal without OPTN and is rescued using a necroptosis inhibitor. These results place OPTN at the crux of neuronal survival from potentially lethal CNS viral infections.

Extradural lumbar nerve root and ganglion capillary hemangioma: case report.

INTRODUCTION: Extradural spinal hemangiomas without vertebral body or intradural involvement are rare and often misdiagnosed. When present in the lumbar spine, they typically present with radiculopathy and weakness. CT imaging is helpful in assessing for bony involvement and temporal involvement while MRI imaging can be helpful in distinguishing hemangiomas from other mass lesions, however current reports on imaging features are limited. Diagnosis remains primarily dependent on tissue pathology with surgery as the mainstay of treatment. CASE PRESENTATION: We present a unique case report in which we obtain additional DSA imaging to not only visualize the vascular anatomy associated with a L4-5 neuroforaminal capillary hemangioma involving the L4 nerve root and ganglion, but to also embolize the feeding artery prior to surgical resection. Patient initially underwent a CT-guided biopsy that was suspicious for a hemangioma and confirmed on final pathology to be the capillary subtype. DISCUSSION: When diagnosed appropriately, patients avoid unnecessary additional testing and avoid erroneous treatment of this rare lesion. Based on our experience, we propose initial MRI imaging to characterize the hemangioma and evaluate for intradural involvement, DSA to assess vascularity followed by embolization of the lesion when able in order to minimize intra-operative hemorrhage risk, and ultimately surgery to achieve a gross total resection.

Case series of diffuse extraneural metastasis in H3F3A mutant high-grade gliomas: Clinical, molecular phenotype and literature review.

H3K27M and H3.3G34R/V mutations have been identified in pediatric high-grade gliomas (pHGG), though extraneural metastases are rarely reported and poorly characterized. Three pHGG patients from two institutions were identified with extraneural metastasis, harboring histone mutations. Their clinical, imaging and molecular characteristics are reported here. A 17-year old female presented with supratentorial H3.3G34R-mutant glioma with metastatic osseous lesions in the spine, pelvis, bone marrow, pleural effusion and soft tissue of pelvis. Bone marrow biopsy and soft tissue of pelvis biopsy showed neoplastic cells positive for P53. A 20-year old female was diagnosed with H3F3A H3K27M-mutant thalamic glioma. She developed diffuse sclerotic osseous lesions. Biopsy of an osseous lesion was non-diagnostic. A 17-year old female presented with a H3F3A H3K27M-mutant diffuse midline glioma with diffuse spinal cord metastasis. She further developed multifocal chest lymphadenopathy, pleural effusions, and a soft tissue mass in the abdominal wall. The latter was positive for H3K27M mutation. We present the first case series of pHGG with H3F3A mutation and diffuse extraneural dissemination, describing their clinical and molecular profile.

Selective time-dependent changes in activity and cell-specific gene expression in human postmortem brain.

As a means to understand human neuropsychiatric disorders from human brain samples, we compared the transcription patterns and histological features of postmortem brain to fresh human neocortex isolated immediately following surgical removal. Compared to a number of neuropsychiatric disease-associated postmortem transcriptomes, the fresh human brain transcriptome had an entirely unique transcriptional pattern. To understand this difference, we measured genome-wide transcription as a function of time after fresh tissue removal to mimic the postmortem interval. Within a few hours, a selective reduction in the number of neuronal activity-dependent transcripts occurred with relative preservation of housekeeping genes commonly used as a reference for RNA normalization. Gene clustering indicated a rapid reduction in neuronal gene expression with a reciprocal time-dependent increase in astroglial and microglial gene expression that continued to increase for at least 24 h after tissue resection. Predicted transcriptional changes were confirmed histologically on the same tissue demonstrating that while neurons were degenerating, glial cells underwent an outgrowth of their processes. The rapid loss of neuronal genes and reciprocal expression of glial genes highlights highly dynamic transcriptional and cellular changes that occur during the postmortem interval. Understanding these time-dependent changes in gene expression in post mortem brain samples is critical for the interpretation of research studies on human brain disorders.

Optimization of Viable Glioblastoma Cryopreservation for Establishment of Primary Tumor Cell Cultures.

Background: Technologies related to the establishment of primary tumor cell cultures from solid tumors, including glioblastoma, are increasingly important to oncology research and practice. However, processing of fresh tumor specimens for establishment of primary cultures on the day of surgical collection is logistically difficult. The feasibility of viable cryopreservation of glioblastoma specimens, allowing for primary culture establishment weeks to months after surgical tumor collection and freezing, was demonstrated by Mullins et al. in 2013, with a success rate of 59% that was not significantly lower than that achieved with fresh tumor tissue. However, research targeting optimization of viable glioblastoma cryopreservation protocols for establishment of primary tumor cultures has been limited. Objectives: The objective of this study was to optimize glioblastoma cryopreservation methods for viable cryobanking and to determine if two-dimensional (2D) or three-dimensional (3D) culture conditions were more supportive of glioblastoma growth after thawing of frozen tumor specimens. Methods: Portions of eight human glioblastoma specimens were cryopreserved by four different protocols differing in the time of enzymatic digestion (before or after cryopreservation), and in the type of cryopreservation media (CryoStor CS10 or 10% dimethyl sulfoxide and 90% fetal calf serum). After 1 month, frozen tissues were thawed, enzymatically digested, if not digested before, and used for initiation of 2D or 3D primary tumor cultures to determine viability. Results: Among the tested cryopreservation and culturing protocols, the most efficient combinations of cryopreservation and culture were those associated with the use of CryoStor CS10 cryopreservation medium, enzymatic digestion before freezing, and 2D culturing after thawing with a successful culture rate of 8 out of 8 cases (100%). Two-dimensional cultures were in general more efficient for the support of tumor cell growth after thawing than 3D cultures. Conclusions: This study supports development of evidence-based viable glioblastoma cryopreservation methods for use in glioblastoma biobanking and research.

Identification of a periodontal pathogen and bihormonal cells in pancreatic islets of humans and a mouse model of periodontitis.

Results from epidemiological and prospective studies indicate a close association between periodontitis and diabetes. However the mechanisms by which periodontal pathogens influence the development of prediabetes/diabetes are not clear. We previously reported that oral administration of a periodontal pathogen, Porphyromonas gingivalis (Pg) to WT mice results in insulin resistance, hyperinsulinemia, and glucose intolerance and that Pg translocates to the pancreas. In the current study, we determined the specific localization of Pg in relation to mouse and human pancreatic α- and ß-cells using 3-D confocal and immunofluorescence microscopy and orthogonal analyses. Pg/gingipain is intra- or peri-nuclearly localized primarily in ß-cells in experimental mice and also in human post-mortem pancreatic samples. We also identified bihormonal cells in experimental mice as well as human pancreatic samples. A low percentage of bihormonal cells has intracellular Pg in both humans and experimental mice. Our data show that the number of Pg translocated to the pancreas correlates with the number of bihormonal cells in both mice and humans. Our findings suggest that Pg/gingipain translocates to pancreas, particularly ß-cells in both humans and mice, and this is strongly associated with emergence of bihormonal cells.

Intracranial Leiomyoma Associated with Epstein-Barr Virus: A Cerebellopontine Angle Mass Presenting with Trigeminal Neuralgia.

BACKGROUND: Primary intracranial leiomyoma is a rare smooth muscle tumor often associated with Epstein-Barr virus (EBV), with <30 cases reported worldwide. These tumors commonly occur in patients with immunocompromised status, especially those with human immunodeficiency virus. In the present report, we have described the case of an EBV-associated leiomyoma at the cerebellopontine angle. The patient had presented with trigeminal neuralgia, which, to the best of our knowledge, is the first reported anatomical location and presentation for this tumor type. CASE DESCRIPTION: A 41-year-old male patient had presented with right-sided facial pain in the V1 and V2 dermatomes and previous workup and imaging studies. The patient had undergone treatment of a presumed right-side cerebellopontine angle meningioma as determined by the magnetic resonance imaging characteristics (no biopsy). The patient subsequently underwent right-sided retrosigmoid craniotomy and gross total resection of the tumor. The postoperative period was uneventful with resolution of the trigeminal neuralgia. Histopathologic examination revealed spindle cell neoplasm with histopathologic and immunohistochemical features consistent with leiomyoma. The tumor cells were positive for smooth muscle actin and desmin and were negative for S100, SOX-10, epithelial membrane antigen, glial fibrillary acidic protein, progesterone receptor, CD31, CD34, and E-cadherin. CONCLUSIONS: Primary intracranial leiomyomas are rare tumors associated with EBV infection that occur in immunocompromised patients. These lesions should be considered in the differential diagnosis for patients with known immunocompromised status (e.g., human immunodeficiency virus), and tissue biopsy should be considered.

Metalloporphyrin Pd(T4) Exhibits Oncolytic Activity and Cumulative Effects with 5-ALA Photodynamic Treatment against C918 Cells.

Photodynamic therapy is a non-invasive method where light activates a photosensitizer bound to cancer cells, generating reactive oxygen species and resulting in cell death. This study assessed the oncolytic potential of photodynamic therapy, comparing European Medicines Agency and United States Food and Drug Administration-approved 5-aminolevulinic acid (5-ALA) to a metalloporphyrin, Pd(T4), against a highly invasive uveal melanoma cell line (C918) in two- and three-dimensional models in vitro. Epithelial monolayer studies displayed strong oncolytic effects (>70%) when utilizing Pd(T4) at a fraction of the concentration, and reduced pre-illumination time compared to 5-ALA post-405 nm irradiance. When analyzed at sub-optimal concentrations, application of Pd(T4) and 5-ALA with 405 nm displayed cumulative effects. Lethality from Pd(T4)-photodynamic therapy was maintained within a three-dimensional model, including the more resilient vasculogenic mimicry-forming cells, though at lower rates. At high concentrations, modality of cell death exhibited necrosis partially dependent on reactive oxygen species. However, sub-optimal concentrations of photosensitizer exhibited an apoptotic protein expression profile characterized by increased Bax/Bcl-2 ratio and endoplasmic stress-related proteins, along with downregulation of apoptotic inhibitors CIAP-1 and -2. Together, our results indicate Pd(T4) as a strong photosensitizer alone and in combination with 5-ALA against C918 cells.

Sporadic Hemangioblastoma of the Cavernous Sinus and Meckel's Cave.

Objectives To describe an extremely rare case of sporadic hemangioblastoma (HB) within the cavernous sinus and Meckel's cave with extension to the cerebellopontine angle (CPA) cistern. Methods A 73-year-old male presented with hearing loss, unilateral ptosis, and facial numbness. Results The imaging showed a complex cystic-solid mass centered at the left cavernous sinus and Meckel's cave with extension to the CPA cistern. Patient underwent retrosigmoid craniectomy for partial resection of the CPA angle component of the mass. Surgical pathology confirmed the diagnosis of HB and patient was scheduled for subsequent radiotherapy of the residual mass. Conclusions We present an exceptional case of supratentorial HB without associated von Hippel-Lindau (VHL) disease, which was predominantly located in the cavernous sinus and Meckel's cave and led to multiple cranial nerve symptoms. We describe imaging characteristics and radiologic-pathologic correlation of this atypically located HB, which can be difficult to consider in the differential diagnosis presurgically.

Transcriptional Profiling of Patient Isolates Identifies a Novel TOR/Starvation Regulatory Pathway in Cryptococcal Virulence.

Human infection with Cryptococcus causes up to a quarter of a million AIDS-related deaths annually and is the most common cause of nonviral meningitis in the United States. As an opportunistic fungal pathogen, Cryptococcus neoformans is distinguished by its ability to adapt to diverse host environments, including plants, amoebae, and mammals. In the present study, comparative transcriptomics of the fungus within human cerebrospinal fluid identified expression profiles representative of low-nutrient adaptive responses. Transcriptomics of fungal isolates from a cohort of HIV/AIDS patients identified high expression levels of an alternative carbon nutrient transporter gene, STL1, to be associated with poor early fungicidal activity, an important clinical prognostic marker. Mouse modeling and pathway analysis demonstrated a role for STL1 in mammalian pathogenesis and revealed that STL1 expression is regulated by a novel multigene regulatory mechanism involving the CAC2 subunit of the chromatin assembly complex 1, CAF-1. In this pathway, the global regulator of virulence gene VAD1 was found to transcriptionally regulate a cryptococcal homolog of a cytosolic protein, Ecm15, in turn required for nuclear transport of the Cac2 protein. Derepression of STL1 by the CAC2-containing CAF-1 complex was mediated by Cac2 and modulated binding and suppression of the STL1 enhancer element. Derepression of STL1 resulted in enhanced survival and growth of the fungus in the presence of low-nutrient, alternative carbon sources, facilitating virulence in mice. This study underscores the utility of ex vivo expression profiling of fungal clinical isolates and provides fundamental genetic understanding of saprophyte adaption to the human host.IMPORTANCECryptococcus is a fungal pathogen that kills an estimated quarter of a million individuals yearly and is the most common cause of nonviral meningitis in the United States. The fungus is carried in about 10% of the adult population and, after reactivation, causes disease in a wide variety of immunosuppressed individuals, including the HIV infected and patients receiving transplant conditioning, cancer therapy, or corticosteroid therapy for autoimmune diseases. The fungus is widely carried in the soil but can also cause infections in plants and mammals. However, the mechanisms for this widespread ability to infect a variety of hosts are poorly understood. The present study identified adaptation to low nutrients as a key property that allows the fungus to inhabit these diverse environments. Further studies identified a nutrient transporter gene, STL1, to be upregulated under low nutrients and to be associated with early fungicidal activity, a marker of poor clinical outcome in a cohort of HIV/AIDS patients. Understanding molecular mechanisms involved in adaptation to the human host may help to design better methods of control and treatment of widely dispersed fungal pathogens such as Cryptococcus.

Herpes Simplex Virus 1 Infection Promotes the Growth of a Subpopulation of Tumor Cells in Three-Dimensional Uveal Melanoma Cultures.

Herpes simplex virus 1 (HSV-1)-mediated oncolytic therapy is an emerging cancer treatment modality with potential effectiveness against a variety of malignancies. To better understand the interaction of HSV-1 with neoplastic cells, we inoculated three-dimensional (3D) cultures of human uveal melanoma cells with HSV-1. 3D melanoma cultures were established by placing tumor cells on the surface of a Matrigel matrix, which was followed by the growth of tumor cells on the matrix surface and invasion of the Matrigel matrix by some tumor cells to form multicellular tumor spheroids within the matrix. When established 3D melanoma cultures were inoculated with HSV-1 by placing virus on the surface of cultures, virus infection caused extensive death of melanoma cells growing on the surface of the 3D matrix and significantly decreased the number of tumor cell spheroids within the matrix. However, HSV-1 infection did not lead to a complete destruction of tumor cells in the 3D cultures during a 17-day observation period and, surprisingly, HSV-1 infection promoted the growth of some melanoma cells within the matrix as determined by the significantly increased size of residual viable multicellular tumor spheroids in virus-inoculated 3D cultures at 17 days after virus inoculation. Acyclovir treatment inhibited HSV-1-induced tumor cell killing but did not block the virus infection-induced increase in spheroid size. These findings suggest that although HSV-1 oncolytic virotherapy may cause extensive tumor cell killing, it may also be associated with the unintended promotion of the growth of some tumor cells.IMPORTANCE Cancer cells are exposed to HSV-1 during oncolytic virotherapy with the intention of killing tumor cells. Our observations reported here suggest that potential dangers of HSV-1 oncolytic therapy include promotion of growth of some tumor cells. Furthermore, our findings raise the possibility that HSV-1 infection of neoplastic cells during natural infections or vaccinations may promote the growth of tumors. Our study indicates that HSV-1 infection of 3D tumor cell cultures provides an experimental platform in which mechanisms of HSV-1-mediated promotion of tumor cell growth can be effectively studied.

Herpes Simplex Virus Type 1 Human Cervical Dorsal Root Ganglionitis.

We present a case of a 34-year-old right-handed Caucasian male with chronic occipital neuralgia refractory to medical therapies and minimally invasive pain procedures who underwent surgical cervical dorsal root ganglionectomy which completely relieved his headaches. The histopathological and immunohistochemical findings of the resected cervical dorsal root ganglia were consistent with active herpes simplex virus type 1 (HSV-1) infection causing ganglionitis. To the best of our knowledge, this case represents the first histopathologically proven HSV-1 cervical dorsal root ganglionitis in humans. This case provides an insight into a possible etiology of occipital neuralgia.

EBV-positive post-transplant lymphoproliferative disorder presenting as primary diffuse large B-cell lymphoma of the central nervous system.

Primary central nervous system post-transplant lymphoproliferative disorder (PCNS-PTLD) is a rare complication with inferior survival outcomes in solid organ transplant patients. It represents approximately 7-15% of all PTLD patients. Because of the rarity of this disease, the diagnosis of PCNS-PTLD is often challenging, and the optimal therapy has not been established. We report a case of a renal transplant patient who initially presented with acute altered neurological function, an enhancing mass lesion of the brain on magnetic resonance imaging (MRI), and nonspecific reactive histopathological changes on brain biopsy. The lesion was self-limited and spontaneously resolved without medical treatment for PTLD. Six months later, surveillance MRI revealed recurrence of the brain lesion. The biopsy showed morphologic changes consistent with Epstein-Barr virus (EBV)-positive diffuse large B-cell lymphoma. The patient responded well to reduction of immunosuppression and treatment with a single-agent regimen of rituximab. This is an unusual case of PCNS-PTLD with an initial presentation resembling a self-limited reactive lesion.

Multiple cerebral fusiform aneurysms involving the posterior and anterior circulation including the anterior cerebral artery: a case report.

Multiple fusiform aneurysms occurring in a single patient are very uncommon. Fusiform aneurysms affecting the anterior circulation are extremely rare and reported cases involve the middle cerebral artery and internal carotid artery. We report here a case of a 55-year-old African American male who had a history of multiple strokes and who was found unresponsive prior to his latest hospital admission. Neuroimaging was remarkable for multiple fusiform aneurysms involving segments of the anterior, middle and posterior cerebral arteries as well as subarachnoid and intraparenchymal hemorrhage. Unfortunately even after aggressive medical management, the patient did not show any signs of recovery and further management entailed comfort care measures only. The patient passed away shortly thereafter. Post-mortem examination confirmed the presence of multiple fusiform aneurysms with involvement of both the posterior and anterior circulation including the anterior cerebral artery as well as subarachnoid and intraparenchymal hemorrhage. Interestingly, there was no apparent site of aneurysm rupture identified to explain the hemorrhage. Our case is unusual in that the multiple fusiform aneurysms were found to extensively involve both the anterior and posterior circulation. Furthermore, our case demonstrates involvement of the anterior cerebral artery by fusiform aneurysms, which is also a very rare event. .

An Engineered Herpesvirus Activates Dendritic Cells and Induces Protective Immunity.

Herpes simplex viruses (HSV) are human pathogens that switch between lytic and latent infection. While attenuated HSV is explored for vaccine, the underlying event remains poorly defined. Here we report that recombinant HSV-1 with a mutation in the γ134.5 protein, a virulence factor, stimulates dendritic cell (DC) maturation which is dependent on TANK-binding kinase 1 (TBK1). When exposed to CD11+ DCs, the mutant virus that lacks the amino terminus of γ134.5 undergoes temporal replication without production of infectious virus. Mechanistically, this leads to sequential phosphorylation of interferon regulatory factor 3 (IRF3) and p65/RelA. In correlation, DCs up-regulate the expression of co-stimulatory molecules and cytokines. However, selective inhibition of TBK1 precludes phosphorylation of IRF3 and subsequent DC activation by the γ134.5 mutant. Herein, the γ134.5 mutant is immune-stimulatory and non-destructive to DCs. Remarkably, upon immunization the γ134.5 mutant induces protection against lethal challenge by the wild type virus, indicative of its vaccine potential. Furthermore, CD11+ DCs primed by the γ134.5 mutant in vivo mediate protection upon adoptive transfer. These results suggest that activation of TBK1 by engineered HSV is crucial for DC maturation, which may contribute to protective immunity.