A prospective study on the efficacy of oral estrogen in female patients with acromegaly.

PURPOSE: To evaluate the efficacy and safety of oral estrogen therapy in female patients of childbearing age with uncontrolled acromegaly and to verify the significance of the presence of estrogen receptor α (ER-α) in somatotropinomas. METHODS: Prospective study in which biochemical and radiological evaluations were performed at baseline and after six months of treatment with an oral formulation of ethinyl-estradiol 0.03 mg and levonorgestrel 0.15 mg. ER-α was assessed by immunohistochemistry and immunopositivity was considered when it was present in ≥ 1% of cells. RESULTS: Eight patients with uncontrolled acromegaly were selected. All patients underwent surgery. Four patients were on octreotide LAR 30 mg, two patients were on lanreotide autogel 120 mg, and two patients had active disease after surgery. At the end of follow-up, IGF-I normalized in 3/8 (37%), 2/8 (25%) patients presented with mean IGF-I reduction of 25% but without IGF-I normalization, and 2/8 (25%) did not respond-one had a 13% increase in IGF-I and IGF-I level remained unchanged after treatment in the other. In one patient, treatment was discontinued after 3 months due to side effects (headache), with an IGF-I reduction of 28% but without normalization. Tumor volume increase (41%) was observed in only one patient (the only tumor with positive ER-α expression). CONCLUSIONS: In uncontrolled patients with acromegaly, a trial with oral estrogen can be an option for young women. Oral estrogen was well tolerated, but the somatotropinoma that presented ER-α expression was the only somatotropinoma that presented growth during treatment.

Congenital Zika syndrome is associated with maternal protein malnutrition.

Zika virus (ZIKV) infection during pregnancy is associated with a spectrum of developmental impairments known as congenital Zika syndrome (CZS). The prevalence of this syndrome varies across ZIKV endemic regions, suggesting that its occurrence could depend on cofactors. Here, we evaluate the relevance of protein malnutrition for the emergence of CZS. Epidemiological data from the ZIKV outbreak in the Americas suggest a relationship between undernutrition and cases of microcephaly. To experimentally examine this relationship, we use immunocompetent pregnant mice, which were subjected to protein malnutrition and infected with a Brazilian ZIKV strain. We found that the combination of protein restriction and ZIKV infection leads to severe alterations of placental structure and embryonic body growth, with offspring displaying a reduction in neurogenesis and postnatal brain size. RNA-seq analysis reveals gene expression deregulation required for brain development in infected low-protein progeny. These results suggest that maternal protein malnutrition increases susceptibility to CZS.

Congenital Zika virus infection: a neuropathological review.

BACKGROUND: A relationship between Zika virus (ZikV) infection in pregnancy and the occurrence of microcephaly was established during the Zika outbreak in Brazil (2015-2016). Neuropathological findings in congenital Zika syndrome helped to understand its pathogenetic mechanisms. RESULTS: The most relevant postmortem findings in the central nervous system (CNS) of fetuses and neonates infected with ZikV early in gestation are microcephaly with ex-vacuo ventriculomegaly and large head circumference associated with obstructive hydrocephalus due to severe midbrain and aqueduct distortion. Babies with severe brain lesions are born with arthrogryposis. Histologically, there is extensive destruction of the hemispheric parenchyma, calcifications, various disturbances of neuronal migration, reactive gliosis, microglial hyperplasia and occasional perivascular cuffs of lymphocytes, also in the meninges. Hypoplastic lesions secondary to the lack of descending nerve fibers include small basis pontis, pyramids and spinal corticospinal tracts. Cerebellar hypoplasia is also common. Severe nerve motor nerve cell loss is observed in the anterior horn of the spinal cord. CONCLUSION: A spectrum of neuropathological changes, from severe microcephaly to obstructive hydrocephalus was observed. The severity of the lesions is directly related to the gestational age, the most severe occurring when the mother is infected in the first trimester. Infection of progenitor cells at the germinal matrix was demonstrated. The lack of spinal motor neurons is responsible for fetal acynesia and consequent arthrogryposis.

Myosin Va is developmentally regulated and expressed in the human cerebellum from birth to old age.

Myosin Va functions as a processive, actin-based motor molecule highly enriched in the nervous system, which transports and/or tethers organelles, vesicles, and mRNA and protein translation machinery. Mutation of myosin Va leads to Griscelli disease that is associated with severe neurological deficits and a short life span. Despite playing a critical role in development, the expression of myosin Va in the central nervous system throughout the human life span has not been reported. To address this issue, the cerebellar expression of myosin Va from newborns to elderly humans was studied by immunohistochemistry using an affinity-purified anti-myosin Va antibody. Myosin Va was expressed at all ages from the 10th postnatal day to the 98 th year of life, in molecular, Purkinje and granular cerebellar layers. Cerebellar myosin Va expression did not differ essentially in localization or intensity from childhood to old age, except during the postnatal developmental period. Structures resembling granules and climbing fibers in Purkinje cells were deeply stained. In dentate neurons, long processes were deeply stained by anti-myosin Va, as were punctate nuclear structures. During the first postnatal year, myosin Va was differentially expressed in the external granular layer (EGL). In the EGL, proliferating prospective granule cells were not stained by anti-myosin Va antibody. In contrast, premigratory granule cells in the EGL stained moderately. Granule cells exhibiting a migratory profile in the molecular layer were also moderately stained. In conclusion, neuronal myosin Va is developmentally regulated, and appears to be required for cerebellar function from early postnatal life to senescence.

Myosin Va is developmentally regulated and expressed in the human cerebellum from birth to old age

Myosin Va functions as a processive, actin-based motor molecule highly enriched in the nervous system, which transports and/or tethers organelles, vesicles, and mRNA and protein translation machinery. Mutation of myosin Va leads to Griscelli disease that is associated with severe neurological deficits and a short life span. Despite playing a critical role in development, the expression of myosin Va in the central nervous system throughout the human life span has not been reported. To address this issue, the cerebellar expression of myosin Va from newborns to elderly humans was studied by immunohistochemistry using an affinity-purified anti-myosin Va antibody. Myosin Va was expressed at all ages from the 10th postnatal day to the 98th year of life, in molecular, Purkinje and granular cerebellar layers. Cerebellar myosin Va expression did not differ essentially in localization or intensity from childhood to old age, except during the postnatal developmental period. Structures resembling granules and climbing fibers in Purkinje cells were deeply stained. In dentate neurons, long processes were deeply stained by anti-myosin Va, as were punctate nuclear structures. During the first postnatal year, myosin Va was differentially expressed in the external granular layer (EGL). In the EGL, proliferating prospective granule cells were not stained by anti-myosin Va antibody. In contrast, premigratory granule cells in the EGL stained moderately. Granule cells exhibiting a migratory profile in the molecular layer were also moderately stained. In conclusion, neuronal myosin Va is developmentally regulated, and appears to be required for cerebellar function from early postnatal life to senescence.

Neuroproteomics: an insight into ALS.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease of unknown aetiology. Diagnosis is made through physical examination, electrophysiological findings, and by excluding other conditions. There is not a single biomarker that concludes the diagnosis. The aim of this study was to investigate differentially expressed proteins in cerebrospinal fluid (CSF) of ALS patients compared to control subjects, with the purpose to identify a panel of possible biomarkers for the disease. The differentially expressed spots/proteins were submitted to two-dimensional (2D) electrophoresis and recognized with matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry. Parkin-like and many iron and zinc binding were some of the proteins found in ALS CSF. Parkin is a ligase involved in ubiquitin-proteasome pathway and mutations in the parkin gene are the most common cause of recessive familial Parkinson's disease. Iron and zinc are involved with many important metabolic processes and are related to neurodegenerative disease. Common features of ALS comprise failure of the ubiquitin-proteasome system and increased levels of metal ions in the brain. Therefore, the identification of these proteins can be a significant step in ALS research. These and other identified proteins are discussed in this study.

Co-infection of HIV and tropical infectious agents that affect the nervous system.

Tropical infections refer to a group of diseases usually located in regions with a warm climate, particularly affecting developing countries, partly because of the conditions that allow them to thrive. However, due to the increased international travel, infectious agents that were previously limited to tropical regions pose an increasing threat to populations at risk for opportunistic infection (OI), especially those infected with the HIV. Tropical infections can facilitate HIV transmission and accelerate the progression of asymptomatic HIV infection to AIDS. Some have the potential to alter the epidemiology, natural history, and/or response to treatment of the other. The introduction of highly active antiretroviral therapy has provided a huge benefit for the vast majority of patients infected with the HIV, by allowing the immune system to recover, improving the clinical and radiological results and reducing the number of OI. On the other hand, some patients have developed various disorders of immune reconstitution, resulting in either hyper-immune inflammatory response to an exogenous antigen or autoimmunity. A significant proportion of these cases have been reported in immigrants from tropical countries to high-income countries, therefore awareness of these phenomena is needed since clinical presentations are often atypical and pose diagnostic challenges. This article reviews some of the key diagnostic aspects of tropical infections associated with HIV infection.

Acromegaly secondary to growth hormone-releasing hormone secreted by an incidentally discovered pheochromocytoma.

Ectopic growth hormone-releasing hormone (GHRH)-secreting tumors are rare and cause acromegaly with somatotroph hyperplasia. We report a case of acromegaly secondary to GHRH secretion by an incidentally discovered pheochromocytoma in a normotensive patient. A 23-year-old man presented with signs and symptoms of acromegaly. Laboratory evaluation confirmed the diagnosis and magnetic resonance imaging (MRI) revealed a sellar mass which was thought to be a macroadenoma and surgically resected. The patient was not cured and medical treatment was indicated. An abdominal ultrasound performed before initiation of medical treatment showed a solid/cystic lesion superiorly to the right kidney. An abdominal MRI confirmed an adrenal tumor. Hormonal workup of the adrenal incidentaloma revealed elevated urinary catecholamine and total metanephrines findings strongly suggestive of a pheochromocytoma. Acromegaly was then suspected to be due to ectopic secretion of GHRH by the tumor. Patient underwent surgical resection and histopathologic examination confirmed a pheochromocytoma which stained positively for GHRH. Also, review of the pituitary specimen confirmed somatotrophic hyperplasia. Genetic analysis of the ret proto-oncogene showed no mutation. Pituitary MRI was repeated 10 months after pheochromocytoma resection and revealed a slightly enlarged pituitary and partial empty sella. The diagnosis of acromegaly caused by ectopic production of GHRH is a challenging task. A careful histopathological examination of the surgically excised pituitary tissue has a key role to arouse the suspicion and guide the investigation of a secondary cause of acromegaly.

An immunohistochemical, clinical and electroneuromyographic correlative study of the neural markers in the neuritic form of leprosy.

The nerve biopsies of 11 patients with pure neuritic leprosy were submitted to routine diagnostic procedures and immunoperoxidase staining with antibodies against axonal (neurofilament, nerve growth factor receptor (NGFr), and protein gene product (PGP) 9.5) and Schwann cell (myelin basic protein, S-100 protein, and NGFr) markers. Two pairs of non-adjacent histological cross-sections of the peripheral nerve were removed for quantification. All the fascicles of the nerve were examined with a 10X-ocular and 40X-objective lens. The immunohistochemistry results were compared to the results of semithin section analysis and clinical and electroneuromyographic data. Neurofilament staining was reduced in 100% of the neuritic biopsies. NGFr positivity was also reduced in 81.8%, PGP staining in 100% of the affected nerves, S100 positivity in 90.9%, and myelin basic protein immunoreactivity in 90.9%. Hypoesthesia was associated with decreased NGFr (81.8%) and PGP staining (90.9%). Reduced potential amplitudes (electroneuromyographic data) were found to be associated with reduced PGP 9.5 (63.6%) and nerve fiber neurofilament staining (45.4%) by immunohistochemistry and with loss of myelinated fibers (100%) by semithin section analysis. On the other hand, the small fibers (immunoreactive dots) seen amid inflammatory cells continued to be present even after 40% of the larger myelinated fibers had disappeared. The present study shows an in-depth view of the destructive effects of leprosy upon the expression of neural markers and the integrity of nerve fiber. The association of these structural changes with the clinical and electroneuromyographic manifestations of leprosy peripheral neuropathy was also discussed.

An immunohistochemical, clinical and electroneuromyographic correlative study of the neural markers in the neuritic form of leprosy

The nerve biopsies of 11 patients with pure neuritic leprosy were submitted to routine diagnostic procedures and immunoperoxidase staining with antibodies against axonal (neurofilament, nerve growth factor receptor (NGFr), and protein gene product (PGP) 9.5) and Schwann cell (myelin basic protein, S-100 protein, and NGFr) markers. Two pairs of non-adjacent histological cross-sections of the peripheral nerve were removed for quantification. All the fascicles of the nerve were examined with a 10X-ocular and 40X-objective lens. The immunohistochemistry results were compared to the results of semithin section analysis and clinical and electroneuromyographic data. Neurofilament staining was reduced in 100 percent of the neuritic biopsies. NGFr positivity was also reduced in 81.8 percent, PGP staining in 100 percent of the affected nerves, S100 positivity in 90.9 percent, and myelin basic protein immunoreactivity in 90.9 percent. Hypoesthesia was associated with decreased NGFr (81.8 percent) and PGP staining (90.9 percent). Reduced potential amplitudes (electroneuromyographic data) were found to be associated with reduced PGP 9.5 (63.6 percent) and nerve fiber neurofilament staining (45.4 percent) by immunohistochemistry and with loss of myelinated fibers (100 percent) by semithin section analysis. On the other hand, the small fibers (immunoreactive dots) seen amid inflammatory cells continued to be present even after 40 percent of the larger myelinated fibers had disappeared. The present study shows an in-depth view of the destructive effects of leprosy upon the expression of neural markers and the integrity of nerve fiber. The association of these structural changes with the clinical and electroneuromyographic manifestations of leprosy peripheral neuropathy was also discussed.

Expression of ubiquitin and proteasome in motorneurons and astrocytes of spinal cords from patients with amyotrophic lateral sclerosis.

Proteasome, ubiquitin, GFAP and neurofilament were evaluated in motorneurons and astrocytes of spinal cords of ALS and control cases. ALS neurons exhibited ubiquitin positive inclusions and areas of strong immunoreaction for proteasome. Areas of proteasome stain were observed close to neurofilament positive proximal process enlargement. The percentage of neurons strongly immunoreacted, for proteasome was higher in ALS cases than in controls. Many astrocytes were positive for ubiquitin and proteasome. These results suggest that the ubiquitin-proteasome pathway is involved in the ALS pathogenesis and agree with the view that ALS is a disorder of protein aggregation that affects neurons and nonneuronal cells.

Leprosy: contribution of mast cells to epineurial collagenization.

BACKGROUND: Leprosy, a disease caused by Mycobacterium leprae, is an important health problem worldwide. It is responsible for an irreversible nerve damage in which fibrosis plays an important role. The existence of an interaction between mast cells and different fibrotic conditions has long been observed. Tryptase, the most abundant protein product of human mast cells, has been shown to be mitogenic for fibroblasts and to increase type I collagen production. PATIENTS AND METHODS: In order to explore the possible relationship between tryptase-rich mast cells and nerve fibrosis in leprosy, we studied 24 sural nerve biopsies of patients with leprous neuropathy. Mast cells stained with mouse antihuman mast cell antitryptase clone AA1 as well as fibrosis, were quantitatively estimated in both epi- and endoneurial compartments. RESULTS: There was a remarkable association between collagen increase and tryptase-rich mast cell density in the epineurium but not in the endoneurium of leprous nerves. CONCLUSION: Since the epineurium in leprosy is type I collagen rich, the present findings support a tryptase-rich mast cell contribution to epineurial collagenization in leprosy through their tryptase secretion.

Quantitative evidence for neurofilament heavy subunit aggregation in motor neurons of spinal cords of patients with amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS), a neurodegenerative disease of unknown etiology, affects motor neurons leading to atrophy of skeletal muscles, paralysis and death. There is evidence for the accumulation of neurofilaments (NF) in motor neurons of the spinal cord in ALS cases. NF are major structural elements of the neuronal cytoskeleton. They play an important role in cell architecture and differentiation and in the determination and maintenance of fiber caliber. They are composed of three different polypeptides: light (NF-L), medium (NF-M) and heavy (NF-H) subunits. In the present study, we performed a morphological and quantitative immunohistochemical analysis to evaluate the accumulation of NF and the presence of each subunit in control and ALS cases. Spinal cords from patients without neurological disease and from ALS patients were obtained at autopsy. In all ALS cases there was a marked loss of motor neurons, besides atrophic neurons and preserved neurons with cytoplasmic inclusions, and extensive gliosis. In control cases, the immunoreaction in the cytoplasm of neurons was weak for phosphorylated NF-H, strong for NF-M and weak for NF-L. In ALS cases, anterior horn neurons showed intense immunoreactivity in focal regions of neuronal perikarya for all subunits, although the difference in the integrated optical density was statistically significant only for NF-H. Furthermore, we also observed dilated axons (spheroids), which were immunopositive for NF-H but negative for NF-M and NF-L. In conclusion, we present qualitative and quantitative evidence of NF-H subunit accumulation in neuronal perikarya and spheroids, which suggests a possible role of this subunit in the pathogenesis of ALS.

Quantitative evidence for neurofilament heavy subunit aggregation in motor neurons of spinal cords of patients with amyotrophic lateral sclerosis

Amyotrophic lateral sclerosis (ALS), a neurodegenerative disease of unknown etiology, affects motor neurons leading to atrophy of skeletal muscles, paralysis and death. There is evidence for the accumulation of neurofilaments (NF) in motor neurons of the spinal cord in ALS cases. NF are major structural elements of the neuronal cytoskeleton. They play an important role in cell architecture and differentiation and in the determination and maintenance of fiber caliber. They are composed of three different polypeptides: light (NF-L), medium (NF-M) and heavy (NF-H) subunits. In the present study, we performed a morphological and quantitative immunohistochemical analysis to evaluate the accumulation of NF and the presence of each subunit in control and ALS cases. Spinal cords from patients without neurological disease and from ALS patients were obtained at autopsy. In all ALS cases there was a marked loss of motor neurons, besides atrophic neurons and preserved neurons with cytoplasmic inclusions, and extensive gliosis. In control cases, the immunoreaction in the cytoplasm of neurons was weak for phosphorylated NF-H, strong for NF-M and weak for NF-L. In ALS cases, anterior horn neurons showed intense immunoreactivity in focal regions of neuronal perikarya for all subunits, although the difference in the integrated optical density was statistically significant only for NF-H. Furthermore, we also observed dilated axons (spheroids), which were immunopositive for NF-H but negative for NF-M and NF-L. In conclusion, we present qualitative and quantitative evidence of NF-H subunit accumulation in neuronal perikarya and spheroids, which suggests a possible role of this subunit in the pathogenesis of ALS.

Effects of chronic heart disease on skeletal muscle fiber size.

Size changes in muscle fibers of subjects with chronic heart disease (CHD) have been reported, although a consensus has not been achieved. The aims of the present study were to investigate a possible association between CHD and fiber size changes in the brachial biceps compared to subjects without heart disease. Forty-six muscle samples were obtained in autopsies of individuals (13 to 84 years) without neuromuscular disorders, 19 (10 males and 9 females) with, and 27 (14 males and 13 females) without CHD. In all cases muscle sections were stained with hematoxylin and eosin and processed for the visualization of myofibrillar ATPase activity. The lesser diameter of type 1 and type 2 fibers was obtained tracing their outlines (at least 150 fibers of each type per sample) onto an image analyzer connected to a computer. The results were analyzed statistically comparing males and females with and without CHD. Type 1 fiber mean lesser diameters were 51.51 and 54.52 microm in males (normal range 34-71 microm) and 45.65 and 55.42 microm in females (normal range 34-65 microm) without and with CHD, respectively; type 2 fibers measured 54.31, 58.23, 41.15, and 49.57 microm, respectively (normal range 36-79 microm for males and 32-59 microm for females). No significant difference in fiber size was detected in 24 males with and without CHD, while in 22 females there was a significant increase in size in those with cardiomyopathy. We concluded that CHD does not determine significant changes in fiber size. However, in females, there is some hypertrophy which, despite within normal range, may reflect morphologic heterogeneity of the sample, or the daily life activities in the upper limbs as a compensatory mechanism to fatigability that affect predominantly the lower limbs in subjects with CHD.

Effects of chronic heart disease on skeletal muscle fiber size

Size changes in muscle fibers of subjects with chronic heart disease (CHD) have been reported, although a consensus has not been achieved. The aims of the present study were to investigate a possible association between CHD and fiber size changes in the brachial biceps compared to subjects without heart disease. Forty-six muscle samples were obtained in autopsies of individuals (13 to 84 years) without neuromuscular disorders, 19 (10 males and 9 females) with, and 27 (14 males and 13 females) without CHD. In all cases muscle sections were stained with hematoxylin and eosin and processed for the visualization of myofibrillar ATPase activity. The lesser diameter of type 1 and type 2 fibers was obtained tracing their outlines (at least 150 fibers of each type per sample) onto an image analyzer connected to a computer. The results were analyzed statistically comparing males and females with and without CHD. Type 1 fiber mean lesser diameters were 51.51 and 54.52 æm in males (normal range 34-71 æm) and 45.65 and 55.42 æm in females (normal range 34-65 æm) without and with CHD, respectively; type 2 fibers measured 54.31, 58.23, 41.15, and 49.57 æm, respectively (normal range 36-79 æm for males and 32-59 æm for females). No significant difference in fiber size was detected in 24 males with and without CHD, while in 22 females there was a significant increase in size in those with cardiomyopathy. We concluded that CHD does not determine significant changes in fiber size. However, in females, there is some hypertrophy which, despite within normal range, may reflect morphologic heterogeneity of the sample, or the daily life activities in the upper limbs as a compensatory mechanism to fatigability that affect predominantly the lower limbs in subjects with CHD.

MIB-1 labeling index in astrocytic tumors--a clinicopathologic study.

BACKGROUND: Although neuroimage and surgical techniques have improved substantially, the prognosis of patients with astrocytic tumors remains unchanged. The purpose of this study was to evaluate the proliferative activity in astrocytic tumors in different grades of malignancy and correlate it to other clinical features. PATIENTS AND METHODS: From archival paraffin-embedded surgical specimens of 40 patients of the Ribeirão Preto Medical School with World Health Organization grade II (n = 10), grade III (n = 5) and grade IV astrocytomas (n = 25), the MIB-1 labeling index (LI) was determined using at least a half of the blocks per case. The results were correlated to the biological behavior of the tumors. The aims of this study were to determine the level of MIB-1 LI values (cut-off) that reflect differences in biological behavior of these tumors, the impact on survival of clinical features such as age, tumor location or extension of surgical removal as well as the adjuvant therapy. RESULTS AND CONCLUSIONS: As expected, a wide range of MIB-1 LI values was disclosed (mean of 2.35% in grade II astrocytomas to 12.28% in glioblastomas). A close relationship was found between MIB-1 LI and survival of patients with astrocytomas according to the histological grade. All but 1 recurrent tumor presented higher MIB-1 LI in the second biopsy, and the mean MIB-1 LI of the patients who died in the immediate postoperative period (n = 7) was higher in comparison to the MIB-1 LI of the respective grade. Postoperative radiation therapy was an important factor that affected the survival of patients with high-grade astrocytomas (p = 0.006). MIB-1 LI cut-off of 3% divided the astrocytomas in 2 groups with significantly different survival (p < 0.001): median survival time of 12 months (low-grade) versus 45 months (high-grade). On the other hand, univariate analysis did not show any correlation between survival and extension of surgical resection (radical versus partial), tumor's location or patient's age at surgery.