A Preliminary Investigation into Search and Matching for Tumor Discrimination in World Health Organization Breast Taxonomy Using Deep Networks.

Breast cancer is one of the most common cancers affecting women worldwide. It includes a group of malignant neoplasms with a variety of biological, clinical, and histopathologic characteristics. There are more than 35 different histologic forms of breast lesions that can be classified and diagnosed histologically according to cell morphology, growth, and architecture patterns. Recently, deep learning, in the field of artificial intelligence, has drawn a lot of attention for the computerized representation of medical images. Searchable digital atlases can provide pathologists with patch-matching tools, allowing them to search among evidently diagnosed and treated archival cases, a technology that may be regarded as computational second opinion. In this study, we indexed and analyzed the World Health Organization breast taxonomy (Classification of Tumors fifth ed.) spanning 35 tumor types. We visualized all tumor types using deep features extracted from a state-of-the-art deep-learning model, pretrained on millions of diagnostic histopathology images from the Cancer Genome Atlas repository. Furthermore, we tested the concept of a digital "atlas" as a reference for search and matching with rare test cases. The patch similarity search within the World Health Organization breast taxonomy data reached >88% accuracy when validating through "majority vote" and >91% accuracy when validating using top n tumor types. These results show for the first time that complex relationships among common and rare breast lesions can be investigated using an indexed digital archive.

M918V RET mutation causes familial medullary thyroid carcinoma: study of 8 affected kindreds.

Germline mutations in codon 918 of exon 16 of the RET gene (M918T) are classically associated with multiple endocrine neoplasia type 2B (MEN 2B) with highly aggressive medullary thyroid cancer (MTC), pheochromocytoma and a unique phenotype. The objectives of this study are to describe the rare M918V RET mutation discovered in 8 MTC kindreds from Brazil lacking the MEN 2B phenotype classically observed in M918T patients and to investigate the presence of a founder effect for this germline mutation. Eight apparently sporadic MTC cases were diagnosed with the germline M918V RET mutation. Subsequently, their relatives underwent clinical and genetic assessment (n = 113), and M918V was found in 42 of them. Until today, 20/50 M918V carriers underwent thyroidectomy and all presented MTC/C-cell hyperplasia; the remainder carriers are on clinical follow-up. None of the M918V carriers presented clinical features of MEN 2B. Their clinical presentation was heterogeneous, and the age at tumor diagnosis ranged from 24 to 59 years. Lymph node metastases were present in 12/20 patients, and presumable distant metastases in 2/20; in contrast, we observed a carrier of up to 87 years of age without evidence of MTC. Ethnographic fieldwork and haplotype analyses suggested that the founder mutation first settled in that area fifteen generations ago and originated from Portugal. Our study is the first to demonstrate the RET M918V mutation co-segregating in 8 familial MTC kindreds with validated evidence of a founder effect. We suggest that M918V MTC should be clinically considered an American Thyroid Association (ATA) moderate-risk category.

Integration of a postoperative calcitonin measurement into an anatomical staging system improves initial risk stratification in medullary thyroid cancer.

OBJECTIVE: Staging systems applied to medullary thyroid cancer (MTC) rely on initial clinical and pathological features and do not consider the response to treatment. To determine whether MTC staging can be improved by incorporating the first postoperative calcitonin measurement. PATIENTS AND MEASUREMENTS: Eighty-five patients being monitored for MTC (median follow-up 5 years) were retrospectively classified according to both the American Joint Committee on Cancer (AJCC) and the proposed combined risk stratification system (low, intermediate and high risk), which incorporates the first postoperative calcitonin measurement, using the outcomes no evidence of disease (NED), biochemical evidence of disease, structurally identifiable disease and death. RESULTS: Ninety per cent of AJCC I patients were classified as NED at final follow-up. When we added a postoperative calcitonin measurement, 95% low-risk patients were classified as NED at final follow-up. AJCC stages I and IV were associated, respectively, with no occurrence and a high rate (63%) of structurally identifiable disease. Stages II and III yielded similar predictions of structurally identifiable disease, 13% and 14%, respectively. When we included the postoperative calcitonin level, the patients with structural evidence of disease included none from the low-risk group, 10% from the intermediate group and 63% from the high-risk group. The proportion of variance explained analysis (PVE) was better for the combined risk stratification system (54%) than for the AJCC system alone (32%). CONCLUSION: Including the first postoperative calcitonin measurement with the anatomical staging system can better predict the clinical outcome of patients with MTC and refine the follow-up of these patients.

Development and application of a novel sensitive immunometric assay for calcitonin in a large cohort of patients with medullary and differentiated thyroid cancer, thyroid nodules, and autoimmune thyroid diseases.

BACKGROUND: Serum calcitonin (sCT) is a useful biomarker for medullary thyroid cancer (MTC). Consensus has not been reached concerning sCT measurements in the evaluation of nodular thyroid disease (NTD). OBJECTIVE AND METHODS: We developed a new immunofluorometric assay for sCT and have validated it in samples from 794 patients [203 with MTC, 205 with autoimmune thyroid disease (ATD), 248 with NTD, 80 with differentiated thyroid cancer (DTC) 'free of disease', 58 with chronic renal failure (CRF)] and 178 normal individuals, including samples after pentagastrin tests and samples from the washout of 92 FNA procedures in patients with NTD or MTC. We also compared some samples from patients with low or high calcitonin levels using both this assay and the Nichols Institute Diagnostics (NID) assay. RESULTS: The assay's analytical sensitivity was 1.0 pg/ml. Considering MTC patients prior to surgery, the cut-off values for the 95% reference range were 11.1 pg/ml for males and 5.5 pg/ml for females and employing the ROC curve were 18.4 pg/ml for males and 7.8 pg/ml for females. sCT in patients with MTC was strongly correlated with disease status. Patients with NTD and ATD did not present false-positive results. sCT measurements were significantly correlated with age (excluding MTC and CRF). The NID test had a strong correlation with our assay. A hook effect was observed only with concentrations >200,000 pg/ml. CONCLUSIONS: We developed a novel sCT assay and validated it in healthy subjects, as well as in a large cohort of patients with MTC, NTD, ATD, DTC, and CRF.

Role of dopamine in the recruitment of immune cells to the nigro-striatal dopaminergic structures.

Research indicates that inflammation and microglial activation are involved in the initiation and progression of Parkinson's disease (PD). Neuroinflammation contributes to the infiltration of peripheral immune cells and blood-brain barrier (BBB) leakage, linking peripheral and central inflammatory events in the pathogenesis of PD. Dopamine (DA) likely plays a role in this process. In the present study, the dopaminergic toxin 6-hydroxydopamine (6-OHDA) was used to damage dopaminergic neurons. Injection of 6-OHDA within the nigrostriatal pathway produced loss of astrocytes, disruption of the BBB, microglia activation and a reduction in osteopontin (OPN) immunoreactivity. Depletion of DA content by alpha-methylparatyrosine (α-MPT, a tyrosine hydroxylase inhibitor) reduced the infiltration of peripheral macrophages as well as the 6-OHDA-induced increase in microglial cells. DA could therefore be relevant in sustaining inflammation and lymphocyte recruitment induced by 6-OHDA, supporting DA implication in the degeneration of dopaminergic neurons induced by inflammatory processes.

A ten-year clinical update of a large RET p.Gly533Cys kindred with medullary thyroid carcinoma emphasizes the need for an individualized assessment of affected relatives.

OBJECTIVE: Reviewing the clinical outcomes of a large kindred with a RET p.Gly533Cys mutation, 10 years after the first description of this kindred, has provided an important set of clinical data for healthcare decision-making. DESIGN AND PATIENTS: We identified 728 RET533 Brazilian relatives, spread out over 7 generations. We performed clinical examination, biochemical and imaging analyses in the proband and in 103 carriers. MEASUREMENT AND RESULTS: The proband has been followed without evidence of structural disease in the last 10 years but with elevated calcitonin. The clinical and surgical features of 60 thyroidectomized RET533 relatives were also described. Forty-six patients had MTC (21-72 years), and 11 patients had C-cell hyperplasia (CCH) (5-42 years). Twelve MTC patients with lymph node metastases had a tumour size of 0·7-2·8 cm. Calcitonin level and CEA were correlated with disease stage, and none of the patients presented with an altered PTH or metanephrine. A 63-year-old woman developed pheochromocytoma and breast cancer. Two other RET533 relatives developed lung squamous cell carcinoma and melanoma. CONCLUSIONS: A vast clinical variability in RET533 presentation was observed, ranging from only an elevated calcitonin level (3%) to local metastatic disease (25%). Many individuals were cured (42%) and the majority had controlled chronic disease (56%), reinforcing the need for individualized ongoing risk stratification assessment. The importance of this update relies on the fact that it allows us to delineate the natural history of RET 533 MEN2A 10 years after its first description.

Peripheral inflammation increases the deleterious effect of CNS inflammation on the nigrostriatal dopaminergic system.

Evidence supports the role of inflammation in the development of neurodegenerative diseases. In this work, we are interested in inflammation as a risk factor by itself and not only as a factor contributing to neurodegeneration. We tested the influence of a mild to moderate peripheral inflammation (injection of carrageenan into the paws of rats) on the degeneration of dopaminergic neurons in an animal model based on the intranigral injection of lipopolysaccharide (LPS), a potent inflammatory agent. Overall, the treatment with carrageenan increased the effect of the intranigral injection of LPS on the loss of dopaminergic neurons in the SN along with all the other parameters studied, including: serum levels of the inflammatory markers TNF-α, IL-1ß, IL-6 and C-reactive protein; activation of microglia, expression of proinflammatory cytokines, the adhesion molecule ICAM and the enzyme iNOS, loss of astrocytes and damage to the blood brain barrier (BBB). The possible implication of BBB rupture in the increased loss of dopaminergic neurons has been studied using another Parkinson's disease animal model based on the intraperitoneal injection of rotenone. In this experiment, loss of dopaminergic neurons was also strengthened by carrageenan, without affecting the BBB. In conclusion, our data show that a mild to moderate peripheral inflammation can exacerbate the degeneration of dopaminergic neurons caused by a harmful stimulus.

Ulcerative colitis exacerbates lipopolysaccharide-induced damage to the nigral dopaminergic system: potential risk factor in Parkinson`s disease.

Peripheral inflammation could play a role in the origin and development of certain neurodegenerative disorders. To ascertain this possibility, a model of dopaminergic neurodegeneration based on the injection of the inflammatory agent lipopolysaccharide (LPS) within the substantia nigra was assayed in rats with ulcerative colitis (UC) induced by the ingestion of dextran sulphate sodium. We found an increase in the levels of inflammatory markers from serum (tumor necrosis factor-α, IL-1ß, IL-6 and the acute phase protein C-reactive protein) and substantia nigra (tumor necrosis factor-α, IL-1ß, IL-6, inducible nitric oxide synthase, intercellular adhesion molecule-1, microglial and astroglial populations) of rats with UC, as well as an alteration of the blood-brain barrier permeability and the loss of dopaminergic neurons. UC reinforced the inflammatory and deleterious effects of LPS. On the contrary, clodronate encapsulated in liposomes (ClodLip), which depletes peripheral macrophages, ameliorated the effect of LPS and UC. Peripheral inflammation might represent a risk factor in the development of Parkinson's disease.

[Depression in type 1 diabetes mellitus and associated factors]. / Depresión en la diabetes mellitus tipo 1 y factores asociados.

BACKGROUND AND OBJECTIVE: In recent years, there has been an increased interest in depression and diabetes risk factors. Our objectives were 1) Study the variables associated with the presence of depression in patients with type 1 diabetes mellitus (DM1), 2) to analyze potential risk factors for depression in these patients, and 3) to study a possible explanatory model of depression scores in these patients. PATIENTS AND METHODS: 207 patients with DM1. We evaluated sociodemographic and biomedical variables by means of a structured interview. We assessed psychological variables by means of the Scale for Depression in Type 1 Diabetes (EDDI-1) and the Spanish version of Diabetes Quality of Life (Es DQOL). RESULTS: Prevalence of depression was 21,7%. Variables associated with risk of depression in this sample were to be female; be unemployed; smoking; having complications of diabetes or other physical conditions; not perceiving family support or support from friends or colleagues in relation to diabetes; having a high number of weekly hyperglycemia; and a poor quality of life. A model based on previous research was obtained. This model explains a high percentage of the variability in the scores of patients in the EDDI-1. CONCLUSIONS: These results provide an empirical support to the knowledge of the risk factors associated with depression in patients with DM1. Glycemic control and quality of life have an important effect on the scores of depression in these patients, providing information for their treatment.

Degeneration of dopaminergic neurons induced by thrombin injection in the substantia nigra of the rat is enhanced by dexamethasone: role of monoamine oxidase enzyme.

Anti-inflammatory strategies receive growing attention for their potential to prevent pathological deterioration in disorders such as Parkinson's disease, which is accompanied by inflammatory reactions that might play a critical role in the degeneration of nigral dopaminergic neurons. We investigated the influence of dexamethasone - a potent synthetic member of the glucocorticoids class of steroid hormones that acts as an anti-inflammatory - on the degeneration of the dopaminergic neurons of rats observed after intranigral injection of thrombin, a serine protease that induces inflammation through microglia proliferation and activation. We evaluated tyrosine hydroxylase (TH)-positive neurons as well as astroglial and microglial populations; dexamethasone prevented the loss of astrocytes but was unable to stop microglial proliferation induced by thrombin. Moreover, dexamethasone produced alterations in the levels of nexin and the thrombin receptor PAR-1, and facilitated accumulation of alpha-synuclein induced by thrombin in dopaminergic neurons. Dexamethasone increased oxidative stress and expression of monoamine oxidase A and B, along with changes on different MAP kinases related to degenerative processes, resulting in a bigger loss of dopaminergic neurons after intranigral injection of thrombin in dexamethasone-treated animals. It is interesting to ascertain that inhibition of monoamine oxidase by tranylcypromine prevented neurodegeneration of dopaminergic neurons, thus suggesting that the deleterious effects of dexamethasone might be mediated by monoamine oxidase.

Extensão cranial da veia safena parva: quando o fluxo caudal é normal / Cranial extension of the small saphenous vein: when caudal flow is normal

A extensão cranial da veia safena parva se destaca pelas inúmeras variações anatômicas e diferentes padrões de fluxo que podem ser observados, descritos em trabalhos envolvendo dissecções pós-morte ou cirúrgicas, flebografias e Doppler, que denotam a formação embriológica mais precoce e complexa em relação à safena magna. A observação de um tipo específico de extensão cranial da safena parva onde o sentido das valvas é contrário ao habitualmente observado foi primeiramente caracterizada por Carlo Giacomini, sendo o fluxo caudal nesses casos de aspecto normal sem sinal de incompetência valvar. Este artigo demonstra os padrões anatômicos e de fluxo que podem ser caracterizados na veia safena parva, contribuindo para que aspectos normais do seu fluxo não sejam confundidos com incompetência valvar.

Cranial extension of the small saphenous vein is of special interest due to the number of anatomical variations and different blood flow patterns observed, which have been described in investigations including postmortem or surgical dissections, phlebographies, and Doppler studies, indicating an earlier and more complex embryological development in relation to the great saphenous vein. A specific type of cranial extension of the small saphenous vein, where a reverse flow is often observed in the valves, was first characterized by Carlo Giacomini. In these veins, caudal flow is normal, with no evidence of valvular incompetence. This article describes anatomical and blood flow patterns found in the small saphenous vein, thus contributing to avoid that normal blood flow aspects are misinterpreted as valvular incompetence.

Veias linfonodais: uma causa pouco conhecida de varizes: [revisão] / Lymph node veins: a little-known cause of varicose veins: [review]

As veias linfonodais fazem parte de uma rede venosa no triângulo de Scarpa, que liga em vários pontos o sistema venoso superficial às veias profundas, e podem tanto ser causa de incompetência do sistema venoso superficial quanto estar envolvidas na recorrência pós-safenectomia. Na rotina diária dos exames de Doppler venoso de membros inferiores, temos notado de maneira cada vez mais freqüente a associação das veias linfonodais com varizes primárias e recorrentes. A adequada caracterização dessas veias pode ajudar na compreensão do mecanismo fisiopatológico do aparecimento das varizes e permitir um controle e tratamento mais dirigidos. Este artigo lança luz sobre os aspectos anatômicos e fisiológicos das veias linfonodais, objetivando chamar a atenção dos profissionais envolvidos no diagnóstico de doenças venosas dos membros inferiores para uma causa pouco difundida de varizes.

Lymph node veins are part of a venous network in Scarpa"s triangle, communicating in many points the superficial venous system and the deep veins, and may either be the cause of incompetence of the superficial venous system, or be involved in recurrent varicose veins after saphenous vein stripping. In the daily routine of venous Doppler examination of the lower extremities, an increasingly frequent association of lymph node veins with primary and/or recurrent varicose veins has been noticed. Appropriate characterization of these veins may help to understand the pathophysiological mechanism of varicose vein appearance and provide a more focused approach to follow-up and treatment. This article sheds some light on the anatomical and physiological aspects of lymph node veins, drawing the attention of professionals involved in the diagnosis of venous disorders of the lower extremities to a little-known cause of varicose veins.

The intrastriatal injection of thrombin in rat induced a retrograde apoptotic degeneration of nigral dopaminergic neurons through synaptic elimination.

We have performed intrastriatal injection of thrombin and searched for distant effects in the cell body region. In striatum, thrombin produced a slight loss of striatal neurons as demonstrated by neural nuclei immunostaining - a non-specific neuronal marker - and the expression of glutamic acid decarboxylase 67 mRNA, a specific marker for striatal GABAergic interneurons, the most abundant phenotype in this brain area. Interestingly, striatal neuropil contained many boutons immunostained for synaptic vesicle protein 2 and synaptophysin which colocalize with tyrosine hydroxylase (TH), suggesting a degenerative process with pre-synaptic accumulation of synaptic vesicles. When we studied the effects on substantia nigra, we found the disappearance of dopaminergic neurons, shown by loss of TH immunoreactivity, loss of expression of TH and dopamine transporter mRNAs, and disappearance of FluoroGold-labelled nigral neurons. The degeneration of substantia nigra dopaminergic neurons was produced through up-regulation of cFos mRNA, apoptosis and accumulation of alpha-synuclein shown by colocalization experiments. Thrombin effects could be mediated by protease-activated receptor 4 activation, as protease-activated receptor 4-activating peptide mimicked thrombin effects. Our results point out the possible relationship between synapse elimination and retrograde degeneration in the nigral dopaminergic system.

Intracerebral VEGF injection highly upregulates AQP4 mRNA and protein in the perivascular space and glia limitans externa.

Stroke is the third leading cause of death and the leading cause of adult disability in the industrialized nations. One of the consequences of stroke is blood-brain barrier (BBB) leakage and subsequent edema, which is one of the causes of mortality in this pathology. Aquaporin-4 (AQP4) is the most abundant water channel in the brain. Studies in AQP4 knock-out mice have shown a prominent role of this water channel in edema development and resolution after ischemia. Here we have studied changes in AQP4 mRNA and protein expression in response to vascular endothelial growth factor (VEGF), a potent angiogenic factor. VEGF administration highly upregulated AQP4 mRNA and protein in the ventral midbrain. Perfusion of the animals with FITC-albumin prior to sacrifice demonstrated localization of AQP4 protein in close proximity to the VEGF-induced new blood vessels. Expression levels of AQP4 mRNA were maximum 7 days after VEGF injection whereas our previous report showed that BBB leakage is resolved at this time point. Therefore, we speculate a positive role of AQP4 in edema resolution, which may partially explain the previously reported beneficial effects of delayed VEGF administration in ischemic rats. Our results provide new insights into the molecular changes in the edematous brain and may help in future therapeutical directions.

Simvastatin prevents the inflammatory process and the dopaminergic degeneration induced by the intranigral injection of lipopolysaccharide.

Anti-inflammatory strategies have attracted much interest for their potential to prevent further deterioration of Parkinson's disease. Recent experimental and clinical evidence indicate that statins - extensively used in medical practice as effective lipid-lowering agents - have also anti-inflammatory effects. In this study, we investigated the influence of simvastatin on the degenerative process of the dopaminergic neurons of the rat following intranigral injection of lipopolysaccharide (LPS), a potent inductor of inflammation that we have previously used as an animal model of Parkinson's disease. We evaluated TH positive neurons, astroglial, and microglial populations and found that simvastatin prevented the inflammatory processes, as the induction of interleukin-1beta, tumor necrosis factor-alpha, and iNOS and the consequent dopaminergic degeneration induced by LPS. Moreover, simvastatin produced the activation of the neurotrophic factor BDNF, along with the prevention of the oxidative damage to proteins. Moreover, it also prevents the main changes produced by LPS on different mitogen-activated protein kinases, featured as increases of P-c-Jun N-terminal protein kinase, P-extracellular signal-regulated kinase, p-38, and P-glycogen synthase kinase and the decrease of the promotion of cell survival signals such as cAMP response element-binding protein and Akt. Our results suggest that statins could delay the progression of dopaminergic degeneration in disorders involving inflammatory processes.

Blood-brain barrier disruption induces in vivo degeneration of nigral dopaminergic neurons.

We have evaluated the possibility that changes in the vascular system may constitute a contributing factor for the death of nigral dopaminergic neurons in Parkinson's disease. Thus, we have employed intranigral injections of vascular endothelial growth factor (VEGF), the most potent inducer of blood-brain barrier (BBB) permeability. A single dose of 1 mug of VEGF, chosen from a dose-response study, highly disrupted the BBB in the ventral mesencephalon in a time-dependent manner. A strong regional correlation between BBB disruption and loss of tyrosine hydroxylase-positive neurons was evident. Moreover, Fluoro-Jade B labelling showed the presence of dying neurons in the substantia nigra in response to VEGF injection. High number of TUNEL-positive nuclei was observed in this area along with activation of caspase 3 within nigral dopaminergic neurons. Analysis of the glial population demonstrated a strong inflammatory response and activation of astroglia in response to BBB disruption. We conclude that disruption of the BBB may be a causative factor for degeneration of nigral dopaminergic neurons.

Correlation between circulating biomarkers of oxidative stress of maternal and umbilical cord blood at birth.

The objective of the work was to study the relationship between the oxidative state of the mother and the newborn at the moment of birth. We measured oxidative stress markers (carbonyl groups, lipid peroxides and total antioxidant capacity (TAC)) and found a good correlation between the oxidative state of the normal mother and the neonate, since a high mother oxidative stress corresponds to an even higher oxidative stress of the newborn in umbilical cord blood. We also found that smoking mothers and their newborns had a higher concentration of the carbonyl group, lipid peroxides and less TAC. Newborns from these mothers weighed significantly less than others at birth. These data suggest a need for interest in monitoring the oxidative state of mothers during the pregnancy period, especially taking into account that the oxidative level could be involved in later risks of metabolic diseases for both mother and newborn.

Divergent regulatory mechanisms governing BDNF mRNA expression in cerebral cortex and substantia nigra in response to striatal target ablation.

We have studied the regulation of BDNF mRNA expression in the corticostriatal and nigrostriatal systems following neurotoxin ablation of striatal targets induced by quinolinic acid (QA) or 2S:2'R:3'R:-2-(2'3'-dicarboxycyclopropyl)glycine (DCG-IV) injections. Striatal lesions were verified by quantifying the loss of glutamic acid decarboxylase (GAD) mRNA expression. Levels of BDNF mRNA were analyzed at 6, 12, and 24 h postlesion. Both lesions paradigms highly induced BDNF mRNA in the ipsilateral cerebral cortex at 6 and 12 h postlesion to drop to control levels at 24 h postlesion. These inductions were mostly restricted to cortical layers II/III and V and ipsilateral insular and piriform cortices, which provide the main cortical inputs to the striatum. Analysis of neuronal activation on these areas demonstrated high levels of cFos mRNA in response to the excitotoxic striatal lesions. Dual labeling of cFos and BDNF mRNAs demonstrated a positive correlation between cortical neuronal activation and expression of BDNF mRNA. Consequently, expression of BDNF in cortical areas projecting to striatum is dependent on both target integrity and neuronal activity. Regulation of BNDF mRNA in substantia nigra, the second major source of BDNF to striatal cells, highly differed from that seen in cerebral cortex. Analysis of cellular expression alone or in combination of BDNF, cFos, tyrosine hydroxylase and GAD mRNAs demonstrated that expression of BDNF in substantia nigra is dependent on target integrity and independent of neuronal activity. In addition, we have studied regulatory mechanisms of BDNF mRNA in the subthalamic nucleus.

In vivo expression of aquaporin-4 by reactive microglia.

Aquaporin-4 (AQP4) is the most abundant aquaporin in the brain and it is widely accepted that this AQP is solely expressed by astrocytes and ependymal cells. AQP4 is particularly enriched in plasma membranes of ependymal cells and astrocyte membrane domains facing blood vessels and pia. AQP4 has gained much attraction due to its involvement in the physiopathology of brain edema, a major cause of death in humans. Consequently, it is of paramount importance to ascertain the phenotypic nature of AQP4 mRNA-expressing cells in the CNS before attempting future clinical studies aimed at minimizing the development of brain edema. We have used intranigral injections of lipopolysaccharide (LPS), a potent immunostimulant that causes disruption of the blood brain barrier, vasogenic edema, loss of reactive astrocytes and activation of microglial cells. These LPS-induced features are ideal for testing the possibility that reactive microglial cells express AQP4 in the adult brain. We have studied AQP4 at the mRNA and protein level. We provide strong evidence that reactive microglial cells highly express AQP4 mRNA and protein in response to LPS injections.

Minocycline reduces the lipopolysaccharide-induced inflammatory reaction, peroxynitrite-mediated nitration of proteins, disruption of the blood-brain barrier, and damage in the nigral dopaminergic system.

We have evaluated the potential neuroprotectant activity of minocycline in an animal model of Parkinson's disease induced by intranigral injection of lipopolysaccharide. Minocycline treatment was very effective in protecting number of nigral dopaminergic neurons and loss of reactive astrocytes at 7 days postlesion. Evaluation of microglia revealed that minocycline treatment highly prevented the lipopolysaccharide-induced activation of reactive microglia as visualized by OX-42 and OX-6 immunohistochemistry. Short-term RT-PCR analysis demonstrated that minocycline partially prevented the lipopolysaccharide-induced increases of mRNA levels for interleukin-1alpha and tumor necrosis factor-alpha. In addition, lipopolysaccharide highly induced protein nitration as seen by 3-nitrotyrosine immunoreactivity in the ventral mesencephalon. Minocycline treatment strongly diminished the extent of 3-nitrotyrosine immunoreactivity. We also found a direct correlation between location of IgG immunoreactivity-a marker of blood-brain barrier disruption-and neurodegenerative processes including death of nigral dopaminergic cells and reactive astrocytes. There was also a precise spatial correlation between disruption of blood-brain barrier and 3-nitrotyrosine immunoreactivity. We discuss potential involvement of lipopolysaccharide-induced formation of peroxynitrites and cytokines in the pathological events in substantia nigra in response to inflammation. If inflammation is proved to be involved in the ethiopathology of Parkinson's disease, our data support the use of minocycline in parkinsonian patients.