[Flow cytometry increases the proportion of valuable samples in cerebrospinal fluid with normal cell count in malignant blood diseases]. / La citometría de flujo aumenta la proporción de muestras valorables en líquido cefalorraquídeo con recuento celular normal en hemopatías malignas.

BACKGROUND: The alteration of cerebrospinal fluid (CSF) in hematologic neoplasms is a poor prognostic marker. The characteristics of CSF are usually analyzed by flow cytometry or cytology. However, paucicellular CSF samples (≤5 cells/dL) can sometimes be considered unsuitable for analysis due to the low number of events. OBJECTIVE: To evaluate the proportion of samples reported as suitable for analysis obtained by cytometry (FCM) and cytology in paucicellular CSF samples. MATERIAL AND METHODS: 169 samples ofpaucicellular CSF corresponding to 115 patients with hematologic neoplasms were selected. The samples were obtained by lumbar puncture in tubes conditioned with EDTA and Transfix®. We characterized the immunophenotype ofCSF samples with an 8-color panel, and 55 samples (32%) were in a small sample tube (SST). In all cases, monocytes were identified by CD14 labeling and T lymphocytes by CD3 labeling. The acquisition was carried out in a FACSCantoII® cytometer, and the analysis was performed using Infinicyt® software. RESULTS: The proportion of samples suitable for analysis was higher in FCM compared to cytology (98% vs 61%, p < 0.000). We identified the presence of T lymphocytes and/or monocytes in most samples (98% and 90%, respectively). In the SST samples, the number of events recorded in low-volume samples (< 1 mL) was lower than in samples with higher volume (140 vs 556, p < 0.001), with a median of identification of 3 cell populations. CONCLUSION: FCM allows the analysis of a higher proportion ofpaucicellular CSF samples than cytology in hematologic neoplasms study.

Limbic encephalitis in a child with ovarian teratoma and influenza B. Case report and critical review of the history of autoimmune anti-N-methyl-d-aspartate receptor encephalitis.

We report the appearance of clinical symptoms and signs of N-methyl-d-Aspartate (NMDA) receptor encephalitis in a patient presenting just days after contraction of influenza B. The offending mature ovarian teratoma was identified and removed on the 10th day after the appearance of symptoms, with subsequent nearly complete resolution of symptoms over the subsequent 6 months. We provide a focused literature review of the clinical and pathophysiologic literature of anti-NMDA receptor encephalitis pertaining to influenza B virus and the pediatric population. Taken together, this study contributes to the pathophysiological understanding of anti-NMDA receptor encephalitis and aids clinicians in its early recognition and management.

Immune cell compartmentalization for brain surveillance and protection.

For decades, it was commonly accepted that the brain is secluded from peripheral immune activity and is self-sufficient for its maintenance and repair. This simplistic perception was based on the presence of resident immune cells, the microglia, and barrier systems within the brain, and the assumption that the central nervous system (CNS) lacks lymphatic drainage. This view was revised with the discoveries that higher functions of the CNS, homeostasis and repair are supported by peripheral innate and adaptive immune cells. The findings of bone marrow-derived immune cells in specialized niches, and the renewed observation that a lymphatic drainage system exists within the brain, further contributed to this revised model. In this Review, we describe the immune niches within the brain, the contribution of professional immune cells to brain functions, the bidirectional relationships between the CNS and the immune system and the relevance of immune components to brain aging and neurodegenerative diseases.

Flow cytometric analysis of cerebrospinal fluid improves detection of leukaemic blasts in infants with acute lymphoblastic leukaemia.

Infants with acute lymphoblastic leukaemia (ALL) have a high frequency of central nervous system (CNS) involvement. Flow cytometric analysis of cerebrospinal fluid (CSF) was recently demonstrated to be a sensitive method for detecting CNS involvement in childhood ALL. In the present study, CSF from 14 infants was collected at routine lumbar punctures and analysed by multicolour flow cytometry. At initial diagnosis, leukaemic blasts were detected in CSF by flow cytometry in 11 patients (78·6%) compared to seven patients (50%) by cytospin. Larger studies are needed to determine if CSF flow cytometry has prognostic value in infant ALL.

Automated Analysis of Cerebrospinal Fluid Cells Using Commercially Available Blood Cell Analysis Devices-A Critical Appraisal.

The analysis of cells in the cerebrospinal fluid (CSF) is a routine procedure that is usually performed manually using the Fuchs-Rosenthal chamber and cell microscopy for cell counting and differentiation. In order to reduce the requirement for manual assessment, automated analyses by devices mainly used for blood cell analysis have been also used for CSF samples. Here, we summarize the current state of investigations using these automated devices and critically review their limitations. Despite technical improvements, the lower limit for reliable leukocyte counts in the CSF is still at approximately 20 cells/µL, to be validated depending on the device. Since the critical range for clinical decisions is in the range of 5-30 cells/µL this implies that cell numbers < 30/µL require a manual confirmation. Moreover, the lower limit of reliable erythrocyte detection by automated devices is at approximately 1000/µL. However, even low erythrocyte numbers may be of clinical importance. In contrast, heavily hemorrhagic samples from neurosurgery may be counted automatically at an acceptable precision more quickly. Finally, cell differentiation by automated devices provides only a rough orientation for lymphocytes, granulocytes and monocytes. Other diagnostically important cell types such as tumor cells, siderophages, blasts and others are not reliably detected. Thus, although the automation may give a gross estimate sufficient for the emergency room situation, each CSF requires a manual microscopy for cytological evaluation for the final report. In conclusion, although automated analysis of CSF cells may provide a first orientation of the cell profile in an individual sample, an additional manual cell count and a microscopic cytology are still required and represent the gold standard.

Characteristics of Central Nervous System (CNS) Involvement in Children With Non-Hodgkin's Lymphoma (NHL) and the Diagnostic Value of CSF Flow Cytometry in CNS Positive Disease.

OBJECTIVE: To investigate the characteristics of central nervous system (CNS) involvement in children with non-Hodgkin's lymphoma (NHL) and the value of flow cytometry (FC) in the diagnosis of CNS disease in pediatric NHL. METHODS: The data of 56 newly diagnosed pediatric NHL patients with CNS involvement (CNS+/mass, CNS+/palsy, CNS+/CSF) were analyzed. The proportions and formats of CNS disease in different pathological types were compared. In addition, FC and conventional cytology (CC) of cerebrospinal fluid (CSF) were carried out in 383 newly diagnosed NHL cases. RESULTS: A total of 383 children with NHL were enrolled. Among these patients, 56 (14.6%) were diagnosed with positive CNS involvement (CNS+), 33 had bulky disease (tumor diameter >10 cm), 32 had bone marrow invasion, 32 had lactate dehydrogenase levels >1000 U/L, and 25 had invasion of more than 4 organs at the time of diagnosis. There were 14 patients with T lymphoblastic lymphoma (T-LBL), 9 with B lymphoblastic lymphoma (B-LBL), 26 with Burkitt's lymphoma (BL), and 2 with Epstein-Barr virus-positive diffuse large B cell lymphoma (EBV + DLBCL). Among the 56 CNS+ patients, 35 were CSF-positive (CSF+); there were 2 patients who were CSF+ via CC detection and 35 who were CSF+ via FC detection. The difference between CC and FC was statistically significant (P < 0.01). In the T-LBL group, 14 patients were CNS+/CSF, and in the B-LBL group, 8 were CNS+/mass. In the BL group, 22 patients were CNS+/mass and 15 were CNS+/CSF. In the anaplastic large-cell lymphoma group, 5 patients were CNS+/mass. Nine of the 56 CNS+ patients had events. The 2-year overall survival rate was 87% ± 0.046%, and the 2-year event-free survival rate was 76.2% ± 0.07%. CONCLUSION: CNS+ diagnoses were more common in pediatric NHL patients with bulky disease and/or bone marrow involvement and/or involvement of more than 4 organs at the time of diagnosis, and they were also common in the EBV + DLBCL and BL groups. FC of CSF showed important clinical significance in the diagnosis of CNS disease in pediatric NHL patients, and it can be used to significantly improve the CNS+ detection rate.

Cerebrospinal Fluid Analysis.

Cerebrospinal fluid (CSF) analysis is a diagnostic tool for many conditions affecting the central nervous system. Urgent indications for lumbar puncture include suspected central nervous system infection or subarachnoid hemorrhage. CSF analysis is not necessarily diagnostic but can be useful in the evaluation of other neurologic conditions, such as spontaneous intracranial hypotension, idiopathic intracranial hypertension, multiple sclerosis, Guillain-Barré syndrome, and malignancy. Bacterial meningitis has a high mortality rate and characteristic effects on CSF white blood cell counts, CSF protein levels, and the CSF:serum glucose ratio. CSF culture can identify causative organisms and antibiotic sensitivities. Viral meningitis can present similarly to bacterial meningitis but usually has a low mortality rate. Adjunctive tests such as CSF lactate measurement, latex agglutination, and polymerase chain reaction testing can help differentiate between bacterial and viral causes of meningitis. Immunocompromised patients may have meningitis caused by tuberculosis, neurosyphilis, or fungal or parasitic infections. Subarachnoid hemorrhage has a high mortality rate, and rapid diagnosis is key to improve outcomes. Computed tomography of the head is nearly 100% sensitive for subarachnoid hemorrhage in the first six hours after symptom onset, but CSF analysis may be required if there is a delay in presentation or if imaging findings are equivocal. Xanthochromia and an elevated red blood cell count are characteristic CSF findings in patients with subarachnoid hemorrhage. Leptomeningeal carcinomatosis can mimic central nervous system infection. It has a poor prognosis, and large-volume CSF cytology is diagnostic.

Cerebrospinal fluid analysis in dogs: Main patterns and prevalence of albuminocytological dissociation.

BACKGROUND: Albuminocytological dissociation (ACD) of the cerebrospinal fluid (CSF) is defined as an increased total protein concentration with normal total nucleated cell count. It is suspected to occur in diseases that alter the blood-brain barrier, increase the production of protein or obstruct the flow of CSF. The purposes of this study were to review the CSF analysis results of a large cohort of dogs with neurological conditions, to analyse the total prevalence of ACD and to describe which diseases have a higher prevalence of ACD. STUDY DESIGN AND METHODS: Medical records were retrospectively searched for dogs whom CSF was sampled from 2012-2019. Data collected included signalment, body weight, site of collection of the CSF, CSF analysis results, and final diagnosis. RESULTS: A total of 497 dogs met the inclusion criteria. ACD was identified in 16.5% (82/497) of dogs. The diseases with higher proportion of ACD were cranial nerve neuropathy (6/10; 60.0%), brain tumour (10/24; 41.7%), idiopathic vestibular disease (7/17; 41.2%) and brain vascular disease (4/13; 30.8%). CLINICAL SIGNIFICANCE: This study describes the CSF patterns of the most common neurological conditions in dogs, also characterizing, for the first time in dogs, the prevalence and causes of ACD, which was identified in 16.5% of the samples. The diseases with highest proportions of ACD were cranial nerve neuropathy, brain tumour, idiopathic vestibular disease and brain vascular disease.

Cytologic diagnosis of metastatic embryonal rhabdomyosarcoma in cerebrospinal fluid: A case report.

Rhabdomyosarcoma (RMS) originates from a differentiation block in muscle progenitors. Leptomeningeal metastasis is a rare but devastating complication of RMS which can be caused by dissemination of cancer cells in cerebrospinal fluid (CSF). Here, we present a 4-year-old female with RMS originating from the upper nasal wall. The following histologic and immunohistochemistry analyses combined with molecular testing analysis supported the diagnosis of embryonal rhabdomyosarcoma (ERMS). Results from CSF routine test, magnetic resonance imaging scans and CSF cytology indicated metastatic meningitis, thus confirming the diagnosis of metastatic ERMS in CSF. This is the first report to describe the clinical features of ERMS in CSF.

Standardizing a volume benchmark for cerebrospinal fluids for optimal diagnostic accuracy.

INTRODUCTION: Cerebral spinal fluid (CSF) cytomorphologic analysis remains the gold standard in the evaluation of malignant leptomeningeal involvement. However, collection of optimal volumes for adequate cytomorphologic evaluation is not standardized. Our study investigated optimal CSF volumes that result in a significant diagnostic result. METHODS: A total of 4114 samples were retrospectively identified from 2014 to 2018, and 2557 samples had concurrent flow cytometry (FC) study. Each specimen was grouped as unsatisfactory, negative, atypical, or positive. Positive samples were grouped as either solid tumors, leukemia, or lymphoma by the type of malignancy detected. Demographic data as well as CSF source was recorded. Specimens with FC were separated by detection on cytology and/or FC. A t-test and ANOVA test were used to compare the average volumes for each group. RESULTS: Average volumes for negative, atypical, and positive samples are 7.48 mL (95% CI: 7.33, 7.63), 7.97 mL (95% CI: 7.37, 8.57), and 8.44 mL (95% CI: 7.46, 9.43), respectively. Average volumes for solid tumors, leukemia, and lymphoma positive samples are 12.0 mL (95% CI: 9.11, 14.89), 6.73 mL (95% CI: 5.94, 7.53), and 8.44 mL (95% CI: 6.78, 10.09). For cases with FC, the volumes are 10.11 mL (95% CI: 9.28, 10.96), 7.28 mL (95% CI: 6.87, 7.70), and 6.86 mL (95% CI: 6.25, 7.49) for positive cytology only, positive cytology/FC, and negative for both, respectively. CONCLUSIONS: Our results suggest that higher volumes produce better results for analysis. We recommend an optimal volume of 8.44 mL for cytologic work-up of malignancies. However, optimal volumes may differ based upon malignancy type and utilization of flow cytometry.

Targeted metabolomic profiling of cerebrospinal fluid from patients with progressive multifocal leukoencephalopathy.

Progressive multifocal leukoencephalopathy (PML), caused by JC polyomavirus, is a demyelinating disease of the central nervous system that primarily affects oligodendrocytes. It can cause significant morbidity and mortality. An early diagnosis is of high relevance as timely immune reconstitution is essential. However, diagnosis can be challenging if virus detection via cerebrospinal fluid (CSF) PCR remains negative. Hence, identifying CSF biomarkers for this disease is of crucial importance. We applied a targeted metabolomic screen to CSF from 23 PML patients and eight normal pressure hydrocephalus (NPH) patients as controls. Out of 188 potentially detectable metabolites, 48 (13 amino acids, 4 biogenic amines, 1 acylcarnitine, 21 phosphatidylcholines, 8 sphingolipids, and the sum of hexoses) passed the quality screen and were included in the analyses. Even though there was a tendency towards lower concentrations in PML (mostly of phosphatidylcholines and sphingomyelins), none of the differences between PML and controls in individual metabolite concentrations reached statistical significance (lowest p = 0.104) and there were no potential diagnostic biomarkers (highest area under the ROC curve 0.68). Thus, CSF metabolite changes in PML are likely subtle and possibly larger group sizes and broader metabolite screens are needed to identify potential CSF metabolite biomarkers for PML.

Cerebrospinal fluid analysis in 108 patients with progressive multifocal leukoencephalopathy.

BACKGROUND: Progressive multifocal leukoencephalopathy (PML) is caused by an opportunistic infection with JC polyoma virus (JCPyV) and mainly affects immunocompromised patients. It leads to pronounced demyelination of the central nervous system (CNS) resulting in severe disability or even death. Detection of JCPyV DNA in the cerebrospinal fluid (CSF) is usually accepted as proof for the diagnosis of PML. Routine CSF parameters, like CSF cell count, protein concentration, Qalbumin, or intrathecal immunoglobulin synthesis are mostly considered normal. However, this has not been investigated systematically. METHODS: We analyzed routine CSF parameters in a cohort of 108 PML patients that were treated at four different neurological centers in Germany. The patients exhibited different underlying conditions with natalizumab-treated multiple sclerosis (n = 54) and human immunodeficiency virus (HIV)-infection (n = 25) being the most frequent. The data were collected at the respective centers in accordance with local requirements and then jointly analyzed. The total PML cohort was compared with a control group of patients with normal pressure hydrocephalus (NPH) and idiopathic intracranial hypertension (IIH). Multiple sclerosis and HIV patients were additionally compared with their own non-PML control groups. RESULTS: The PML group showed an elevated cell count (p < 0.001) compared to the control group, however, this effect was mainly driven by HIV-PML patients. This subgroup also demonstrated a significantly higher proportion of patients with a disturbed blood-CSF-barrier function. CONCLUSIONS: This comprehensive, retrospective study on CSF diagnostic analysis in PML patients provides insight into the CSF of those patients. It demonstrates that CSF composition in PML patients may be specific for the underlying condition that predisposes for the development of PML and thus data have to be interpreted in this context.

A Single Injection of an Optimized Adeno-Associated Viral Vector into Cerebrospinal Fluid Corrects Neurological Disease in a Murine Model of GM1 Gangliosidosis.

GM1 gangliosidosis is a rare neurodegenerative lysosomal storage disease caused by loss-of-function mutations in the gene encoding beta-galactosidase (ß-gal). There are no approved treatments for GM1 gangliosidosis. Previous studies in animal models have demonstrated that adeno-associated viral (AAV) vector-mediated gene transfer to the brain can restore ß-gal expression and prevent the onset of neurological signs. We developed an optimized AAV vector expressing human ß-gal and evaluated the efficacy of a single intracerebroventricular injection of this vector into the cerebrospinal fluid (CSF) of a murine disease model. The AAV vector administration into the CSF increased ß-gal activity in the brain, reduced neuronal lysosomal storage lesions, prevented the onset of neurological signs and gait abnormalities, and increased survival. These findings demonstrate the potential therapeutic activity of this vector and support its subsequent development for the treatment of GM1 gangliosidosis.

Creative and Innovative Methods and Techniques for the Challenges in the Management of Adult Craniopharyngioma.

Craniopharyngioma remains a major challenge in daily clinical practice. The pathobiology of the tumor is still elusive, and there are no consensus or treatment guidelines on the optimal management strategy for this relatively rare tumor. However, recent technical and scientific advances, including genomic and radiomic profiling, innovation in surgical approaches, more precise radiotherapy protocols, targeted therapy, and restoration of lost functions all have the potential to significantly improve the outcome of patients with craniopharyngioma in the near future. Although many of these innovative tools in the new armamentarium of the clinician are still in their infancy, they could reduce craniopharyngioma-related morbidity and mortality and improve the patients' quality of life. In this article, we discuss these creative and innovative approaches that may offer solutions to the obstacles faced in treating craniopharyngioma and future possibilities in improving the care of these patients.

Discrepancy between ventricular and lumbar CSF in chronic meningitis.

Meningitis patient can present with various manifestation including hydrocephalus due to multiple reason. Diagnosis of meningitis mainly rely on CSF analysis which is usually obtained from lumbar puncture. In case of hydrocephalus CSF can be obtain from ventricles during VP shunt operation. Sometimes ventricular CSF can be normal in meningitis patient while lumbar CSF shows abnormality. Possible mechanisms behind this phenomenon are discussed here. Patients who present with hydrocephalus and have normal Ventricular CSF should investigated with lumbar CSF analysis in a view of delay in diagnosis and treatment.

Astrocytic IGFBP2 and CHI3L1 in cerebrospinal fluid drive cortical metastasis of HER2+breast cancer.

The brain is often reported as the first site of recurrence among breast cancer patients overexpressing human epidermal growth factor receptor 2 (HER2). Although most HER2+tumors metastasize to the subcortical region of the brain, a subset develops in the cortical region. We hypothesize that factors in cerebrospinal fluid (CSF) play a critical role in the adaptation, proliferation, and establishment of cortical metastases. We established novel cell lines using patient biopsies to model breast cancer cortical and subcortical metastases. We assessed the localization and growth of these cells in vivo and proliferation and apoptosis in vitro under various conditions. Proteomic analysis of human CSF identified astrocyte-derived factors that support the proliferation of cortical metastases, and we used neutralizing antibodies to test the effects of inhibiting these factors both in vivo and in vitro. The cortical breast cancer brain metastatic cells exhibited greater proliferation than subcortical breast cancer brain metastatic cells in CSF containing several growth factors that nourish both the CNS and tumor cells. Specifically, the astrocytic paracrine factors IGFBP2 and CHI3LI promoted the proliferation of cortical metastatic cells and the formation of metastatic lesions. Disruption of these factors suppressed astrocyte-tumor cell interactions in vitro and the growth of cortical tumors in vivo. Our findings suggest that inhibition of IGFBP2 and CHI3LI signaling, in addition to existing treatment modalities, may be an effective therapeutic strategy targeting breast cancer cortical metastasis.