Development of polypyrrole (nano)structures decorated with gold nanoparticles toward immunosensing for COVID-19 serological diagnosis.

The rapid and reliable detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) seroconversion in humans is crucial for suitable infection control. In this sense, many studies have focused on increasing the sensibility, lowering the detection limits and minimizing false negative/positive results. Thus, biosensors based on nanoarchitectures of conducting polymers are promising alternatives to more traditional materials since they can hold improved surface area, higher electrical conductivity and electrochemical activity. In this work, we reported the analytical comparison of two different conducting polymers morphologies for the development of an impedimetric biosensor to monitor SARS-CoV-2 seroconversion in humans. Biosensors based on polypyrrole (PPy), synthesized in both globular and nanotubular (NT) morphology, and gold nanoparticles are reported, using a self-assembly monolayer of 3-mercaptopropionic acid and covalently linked SARS-CoV-2 Nucleocapsid protein. First, the novel hybrid materials were characterized by electron microscopy and electrochemical measurements, and the biosensor step-by-step construction was characterized by electrochemical and spectroscopic techniques. As a proof of concept, the biosensor was used for the impedimetric detection of anti-SARS-CoV-2 Nucleocapsid protein monoclonal antibodies. The results showed a linear response for different antibody concentrations, good sensibility and possibility to quantify 7.442 and 0.4 ng/mL of monoclonal antibody for PPy in the globular and NT morphology, respectively. The PPy-NTs biosensor was able to discriminate serum obtained from COVID-19 positive versus negative clinical samples and is a promising tool for COVID-19 immunodiagnostic, which can contribute to further studies concerning rapid, efficient, and reliable detections.

Subdural Hygroma: A Rare Complication of a Common Brain Malformation.

INTRODUCTION: Arachnoid cysts are a common incidental finding on head imaging. While the natural history of these cysts in poorly described, hemorrhage with subdural hygroma formation is rare. We review the clinical course of a patient who developed a subdural hygroma following trauma. CASE: The patient was a previously healthy 14-month-old male who presented to the Emergency Department with vomiting after a fall and was found to have esotropia without other focal neurological deficits and a CT scan consistent with a subdural cerebrospinal fluid collection with midline shift. The patient was treated conservatively and his symptoms resolved. DISCUSSION: Arachnoid cyst rupture is a rare complication which can lead to increased intracranial pressure with devastating consequences. Clinical manifestation can be similar to that of other intracranial pathologies. Prompt diagnosis is required to avoid life-threatening symptoms. CONCLUSION: Arachnoid cyst rupture should be considered when evaluating patients with non-specific neurological symptoms following trauma.

Differences in fish communities on natural versus artificial temperate reefs, groundfish conservation applications in British Columbia.

Human-made marine habitats such as artificial reefs are used to mitigate marine habitat degradation and aid conservation of species at risk. We used ROV and sonar to survey threatened rockfish (Sebastes spp.) and other groundfish species associated with 18 artificial and natural reefs along the south coast of BC, Canada. Using an information-theoretic approach, we found that community composition significantly differed between natural and artificial reefs. Artificial reefs had high variability in rockfish abundance, some supporting very high or low relative abundance. Natural reefs consistently supported intermediate rockfish abundances. Groundfish diversity was significantly greater on natural reefs than artificial reefs. Depth and relief were significant predictors for both abundance and species richness. Interestingly, rockfish abundance was negatively associated with proximity to nearest rockfish conservation area. This research is a first step in understanding causal mechanisms leading to differences between fish communities on artificial reefs in our study system, and which reef attributes may facilitate successful contributions to conservation. Though artificial reefs show promise in the conservation of some threatened species, the maintenance of diverse fish communities depends on protection of heterogenous natural reef communities.

Minocycline treatment following hypoxic/ischaemic injury attenuates white matter injury in a rodent model of periventricular leucomalacia.

AIMS: Periventricular white matter injury in premature infants occurs following hypoxia/ischaemia and systemic infection, and results in hypomyelination, as well as neuromotor and cognitive deficits later in life. Inflammatory infiltrates are seen within human cerebral white matter from periventricular leucomalacia (PVL) cases. METHODS: In this study, we examine the time course of CD-68+ microglial cell responses relative to cell death within white matter following hypoxia/ischaemia in a rat model of PVL. We also tested the efficacy of the minocycline, an agent that suppresses microglial activation, in this model when administered as a post-insult treatment. RESULTS: We show that preoligodendrocyte injury in the post-natal day 6 begins within 24 h and continues for 48-96 h after hypoxia/ischaemia, and that microglial responses occur primarily over the first 96 h following hypoxia/ischaemia. Minocycline treatment over this 96 h time window following the insult resulted in significant protection against white matter injury, and this effect was concomitant with a reduction in CD-68+ microglial cell numbers. CONCLUSIONS: These results suggest that anti-inflammatory treatments may represent a useful strategy in the treatment of PVL, where clinical conditions would favour a post-insult treatment strategy.

Transsulfuration in mammals: fetal and early development of methionine-activating enzyme and its relation to hormonal influences.

The development of activity of methionine-activating enzyme was studied in four organs of the rat. Three different patterns were observed: (a) in the liver, specific activity began to increase in late fetal life and reached a maximum 2 days after birth; (b) in the small intestine, specific activity began to rise in the 2nd wk after birth and reached a maximum at age 18 days; and (c) in the brain and kidney, specific activity did not change markedly from the earliest stage of fetal development studied to adult life. Hydrocortisone increased hepatic methionine-activating enzyme activity as much as 55% in the young rat. However, adrenalectomy in the newborn rat did not prevent the postnatal rise in hepatic methionine-activating enzyme activity, nor did adrenalectomy at age 10 days prevent the developmental rise of intestinal activity at age 18 days. Conjugated estrogens partially inhibited both the neonatal rise in hepatic methionine-activating enzyme activity and the rise in activity after adrenal steroid injection. Injection of L-methionine did not increase hepatic methionine-activating enzyme activity in the developing or adult rat.

Selective decrease of the viability and the sterol content of proliferating versus quiescent glioma cells exposed to 25-hydroxycholesterol.

The effects of 25-hydroxycholesterol (25-OHC), a potent inhibitor of sterol synthesis, on the growth, viability, and sterol content of C-6 rat glioma cells have been studied. Suppression of 3-hydroxy-3-methylglutaryl coenzyme A reductase activity and sterol synthesis in cells that were proliferating in medium supplemented with lipoprotein-poor fetal calf serum caused an arrest of growth after 24 hr. Prolonged incubation of serum-supplemented cells with 25-OHC resulted in a loss of morphological integrity and an 80% decline in cell viability, determined by trypan blue dye exclusion. In contrast, C-6 cells that were induced to enter a quiescent state by removal of serum from the medium remained viable and morphologically differentiated in the presence of 25-OHC. Following the addition of whole fetal calf serum to the medium, serum-free cells that had been incubated with 25-OHC for 3 days were able to resume proliferation. the selective killing of proliferating C-6 glioma cells by 25-OHC was correlated with a 45 to 50% decline in the sterol/phospholipid molar ratio, whereas the sterol/phospholipid ratio in the quiescent cells was not affected by 25-OHC. The results suggest that inhibitors of sterol synthesis may have potential as agents that might selectively decrease the growth and viability of glioma cells in the central nervous system without detriment to the normal nondividing neural cells.

Macrophage content of murine sarcomas and carcinomas: associations with tumor growth parameters and tumor radiocurability.

Experiments were designed to investigate whether the tumor-associated macrophage (TAM) content of murine solid tumors correlates with the clonogenic ability of tumor cells to establish s.c. tumors, tumor growth rate, extent of tumor necrosis, tumor metastatic propensity, and tumor radioresponse. Of 13 tumors studied, 6 were sarcomas and 7 were carcinomas; all tumors were of spontaneous origin in C3Hf/Kam mice, with the exception of one sarcoma that was induced by 3-methylcholanthrene. Tumors were growing in the hind thighs of syngeneic mice, and their TAM content was determined when they were 8 mm in diameter. The TAM content varied greatly among tumors, ranging from 9 to 83%. Tumor bearing mice experienced a reduction of 50% or more in the number of peritoneal macrophages, but the degree of reduction was independent of TAM content. A significant negative correlation was noted between TAM content and TD50 values (i.e., the number of tumor cells needed to produce tumors in 50% of injected sites) and between TAM content and the amount of tumor necrosis. Also, an obvious trend toward positive correlation between TAM content and reduced local tumor radiocurability was apparent. No correlation was found between TAM content and tumor growth rate or metastatic spread. TAM from the NFSA sarcoma (a tumor with a low TD50 value, almost without necrosis, and poorly responsive to radiation) stimulated the in vitro growth of NFSA tumor cells. These observations suggest that high TAM content could be conductive to tumor cell proliferation and could be a factor in poor tumor radioresponse.

Increased ultraviolet sensitivity and chromosomal instability related to P53 function in the xeroderma pigmentosum variant.

The xeroderma pigmentosum (XP) variant (XPV) is a form of XP that has normal excision repair but shows defective DNA replication after UV irradiation. In developing various transformed fibroblast cell lines from these patients, we have found that there are significant phenotypic changes in transformed cells that seem to correlate with inactivation of p53. After transformation with SV40, XPV cell lines are only slightly UV sensitive, like their primary counterparts, but their sensitization with caffeine and the induction of sister chromatid exchanges (SCEs) by UV irradiation are greatly enhanced. After transformation by HPV16 E7, which targets the retinoblastoma cell cycle regulatory gene, there is no change in the UV sensitivity of XPV cells; but, when transformed by HPV16 E6 or E6 and E7 combined, there is a large increase in UV sensitivity and in the induction of SCEs. These changes are not associated with any detectable changes in the reactivation of an externally irradiated luciferase expression vector, the excision of cyclobutane pyrimidine dimers from bulk DNA, or unscheduled DNA synthesis and, therefore, do not involve excision repair. We suggest that if SCEs represent homologous recombination between sister chromatids, then in the absence of p53 function, the DNA chain arrest typical of UV-damaged XPV cells initiates strand exchange during recovery. In untransformed cells with normal p53, the preferred mode of recovery would then be replication bypass. The symptoms of elevated solar carcinogenesis in XPV patients may, therefore, be associated with increased genomic instability in cells of the skin in which p53 is inactivated by UV-induced mutations.

NBQX attenuates excitotoxic injury in developing white matter.

The excitatory neurotransmitter glutamate is released from axons and glia under hypoxic/ischemic conditions. In vitro, oligodendrocytes (OLs) express non-NMDA glutamate receptors (GluRs) and are susceptible to GluR-mediated excitotoxicity. We evaluated the role of GluR-mediated OL excitotoxicity in hypoxic/ischemic white matter injury in the developing brain. Hypoxic/ischemic white matter injury is thought to mediate periventricular leukomalacia, an age-dependent white matter lesion seen in preterm infants and a common antecedent to cerebral palsy. Hypoxia/ischemia in rat pups at postnatal day 7 (P7) produced selective white matter lesions and OL death. Furthermore, OLs in pericallosal white matter express non-NMDA GluRs at P7. Unilateral carotid ligation in combination with hypoxia (6% O(2) for 1 hr) resulted in selective, subcortical white matter injury with a marked ipsilateral decrease in immature and myelin basic protein-expressing OLs that was also significantly attenuated by 6-nitro-7-sulfamoylbenzo(f)quinoxaline-2,3-dione (NBQX). Intracerebral AMPA demonstrated greater susceptibility to OL injury at P7 than in younger or older pups, and this was attenuated by systemic pretreatment with the AMPA antagonist NBQX. These results indicate a parallel, maturation-dependent susceptibility of immature OLs to AMPA and hypoxia/ischemia. The protective efficacy of NBQX suggests a role for glutamate receptor-mediated excitotoxic OL injury in immature white matter in vivo.

Late oligodendrocyte progenitors coincide with the developmental window of vulnerability for human perinatal white matter injury.

Hypoxic-ischemic injury to the periventricular cerebral white matter [periventricular leukomalacia (PVL)] results in cerebral palsy and is the leading cause of brain injury in premature infants. The principal feature of PVL is a chronic disturbance of myelination and suggests that oligodendrocyte (OL) lineage progression is disrupted by ischemic injury. We determined the OL lineage stages at risk for injury during the developmental window of vulnerability for PVL (23-32 weeks, postconceptional age). In 26 normal control autopsy human brains, OL lineage progression was defined in parietal white matter, a region of predilection for PVL. Three successive OL stages, the late OL progenitor, the immature OL, and the mature OL, were characterized between 18 and 41 weeks with anti-NG2 proteoglycan, O4, O1, and anti-myelin basic protein (anti-MBP) antibodies. NG2+O4+ late OL progenitors were the predominant stage throughout the latter half of gestation. Between 18 and 27 weeks, O4+O1+ immature OLs were a minor population (9.9 +/- 2.1% of total OLs; n = 9). Between 28 and 41 weeks, an increase in immature OLs to 30.9 +/- 2.1% of total OLs (n = 9) was accompanied by a progressive increase in MBP+ myelin sheaths that were restricted to the periventricular white matter. The developmental window of high risk for PVL thus precedes the onset of myelination and identifies the late OL progenitor as the major potential target. Moreover, the decline in incidence of PVL at approximately 32 weeks coincides with the onset of myelination in the periventricular white matter and suggests that the risk for PVL is related to the presence of late OL progenitors in the periventricular white matter.

A potential role for phospholipids in the regulation of 3-hydroxy-3-methylglutaryl-coenzyme A reductase in cultured C-6 glial cells. Effects of N,N-dimethylethanolamine.

The relation of the activity of the microsomal enzyme, 3-hydroxy-3-methylglutaryl coenzyme A reductase, to cellular phospholipid composition was studied in C-6 glial cells. Phospholipid composition was perturbed by growth of cells in the naturally occurring amino alcohol, N,N-dimethylethanolamine. After growth of C-6 glia in 5 mM N,N-dimethylethanolamine for 24 h, reductase activity was diminished by 50%. A similar diminution in cholesterol synthesis was observed. This effect was not accompanied by any parallel change in cell growth, DNA synthesis, protein synthesis, fatty acid synthetase activity, or microsomal NADPH-cytochrome c reductase activity. The inhibition of reductase activity by N,N-dimethylethanolamine was prevented by the addition of equimolar concentrations of choline to the culture medium and, also, could be reversed completely by removal of N,N-dimethylethanolamine from the culture medium. The effect of N,N-dimethylethanolamine on reductase was associated with the formation of phosphatidyl-N,N-dimethylethanolamine which accumulated primarily at the expense of phosphatidylcholine and, after 24 h, accounted for 27% of total phospholipid phosphorus. The data demonstrate that incorporation of N,N-dimethylethanolamine into the polar head group of cellular phospholipids has a major impact on the regulation of the reductase. These observations may have particular relevance for the mechanisms of regulation of this enzyme, the cellular adaptation to alterations in membrane lipid composition, and the regulation of cholesterol synthesis in the developing nervous system.

Long-term regulation by theophylline of fatty acid synthetase, acetyl-CoA carboxylase and lipid synthesis in cultured glial cells.

The long-term regulation of fatty acid synthetase and acetyl-CoA carboxylase and of fatty acid and sterol synthesis was studied in C-6 glial cells in culture. When theophylline (10(-3) M) was added to the culture medium of these cells, rates of lipid synthesis from acetate and activities of synthetase and carboxylase became distinctly lower than in cells that were untreated. This effect appeared after approximately 12 h, and after 48 h enzymatic activities were reduced approx. 2-fold and rates of lipid synthesis from acetate 3- to 4-fold. The likelihood that the decrease in fatty acid synthesis from acetate was caused by the decrease in activities of fatty acid synthetase and acetyl-CoA carboxylase was established by several observations. These indicated that the locus of the effect probably did not reside at the level of acetate uptake into the cell, alterations in acetate pool sizes or conversion of acetate to acetyl-CoA. Moreover, de novo fatty acid synthesis was found to be the predominant pathway in these glial cells, whether treated with theophylline or not. The mechanism of the effect of theophylline on fatty acid synthetase was shown by immunochemical techniques to involve an alteration in content of enzyme rather than in catalytic efficiency. The change in content of fatty acid synthetase was shown by isotopic-immunochemical experiments to involve a decrease in synthesis of the enzyme. The mechanism whereby theophylline leads to a decrease in lipogenesis and in the synthesis of fatty acid synthetase may not be mediated entirely by inhibition of phosphodiesterase and an increase in cyclic AMP levels, because dibutyryl cyclic AMP (10(-3) M) only partially reproduced the effect.

Regulation of hepatic fatty acid synthetase in the obese-hyperglycemic mutant mouse.

Regulation of fatty acid synthetase has been studied in the obese-hyperglycemic mouse and compared with regulation in non obese, littermate control animals. The mechanisms underlying the regulatory changes were defined by immunochemical techniques. Several major conclusions are justified from the data obtained: (1) Although the hepatic specific activity of fatty acid synthetase is higher in obese than in non obese animals pair-fed chow, no difference in hepatic activities is apparent in animals pair-fed the fat-free diet; (2) The higher enzymatic activity in obese animals fed chow is related to a higher content of enzyme, and this higher content is associated with a higher rate of enzyme synthesis; (3) The decrease in hepatic synthetase activity with starvation is distinctly more striking in non obese than in obese animals, and the changes in activity reflect changes in content of enzyme; (4) With starvation there is a decrease in synthesis of enzyme in obese and non obese animals, but only in non obese animals is there also a marked increase in the rate of synthetase degradation (t1/2 = 24 h during starvation, t1/2 = 76 h during normalfeeding); (5) Refeeding starved mice a fat-free diet results in a more striking increase in hepatic synthetase activity in non obese than in obese animals; (6) Administration of triiodothyronine causes a more marked increase in hepatic synthetase activity in non obese than in obese animals. The data thus define a variety of differences in regulation of hepatic fatty acid synthetase in mutant and normal animals. The roles of enzyme synthesis and degradation in the etiology of these differences are defined, and possible mechanisms underlying regulation of synthetase synthesis and degradation in normal mammalian liver are suggested by the observations.

Hormonal regulation of fatty acid synthetase, acetyl-CoA carboxylase and fatty acid synthesis in mammalian adipose tissue and liver.

The major objectives of this study were to define the roles of adrenal glucocorticoids and glucagon in the long-term regulation of fatty acid synthetase and acetyl-CoA carboxylase of mammalian adipose tissue and liver. Particular emphasis was given to elucidation of the mechanisms whereby these hormones produce their regulatory effects on enzymatic activity. To dissociate mental manipulation, nutritional conditions were ridgidly controlled in the experiments described. Administration of glucocorticoids to adult rats led to a marked reductionin activities of fatty acid synthetase and carboxylase in adipose in adipose tissue but no change occurred in liver. Adrenalectomy produced an increase in activities of these lipogenic enzymes in adipose tissure, but, again, no change was noted in liver. The decrease in enzymatic activities in adipose tissue with glucocorticoid administration correlated well with a decrease in fatty acid synthesis, determined in vivo by the 3-H2O method. The mechanisms whereby glucocorticoids led to a decrease in fatty acid synthetase activity were elucidated by the use of immunochemical techniques. Thus, the decrease in fatty acid synthetase activity observed in adipose tissue was shown to reflect a decrease in content of enzyme, and not a change in catalytic efficiency. The mechanism underlying the decrease in enzyme content is a decrease in synthesis of the enzyme. The relation of the effects of glucocorticoids to the effects of certain other hormones involved in regulation of lipogenesis was investigated in hypophysectomized and in diabetic animals. Thus, the observation that the glucocorticoid effect on synthetase and carboxylase occurred in adipose tissue of hypophysectomized rats indicated that alterations in levels of other pituitary-regulated hormones were not necessary for the effect. That glucocorticoids play some role in regulation of synthetase and carboxylase in liver, at lease in the diabetic state, was shown by the observation that the low activities of these enzymes in diabetic animals could be restored to normal by adrenalectomy. An even more pronounced restorative effect was apparent in adipose tissue of adrenalectomized, diabetic animals. Administration of glucagon during the refeeding of starved rats resulted in a marked reduction in the induction of fatty acid synthetase, acetyl-CoA carboxylase and in the rate of incorporation of 3-H from 3-H2O into fatty acids in liver, but no change in these parameters occurred in adipose tissue. Administration of theophylline resulted in intermediate reduction in liver. The mechanisms whereby glucagon led tto a decrease in fatty acid synthetase activity were elucidated by the use of immunochemical techniques. Thus, the changes in fatty acid synthetase activity were shown to reflect reductions in content of enzyme. The mechanism underlying these reductions in content is reduced synthesis of enzyme.