Isolated pituitary fossa metastasis from a primary tonsillar squamous cell carcinoma: case report.

OBJECTIVE: This paper presents a case of an isolated pituitary fossa metastasis on a background of a previously treated tonsillar squamous cell carcinoma. CASE REPORT: A 64-year-old male, diagnosed with a primary p16-negative squamous cell carcinoma in the right tonsil, was treated with a course of chemoradiotherapy with curative intent. Positron emission tomography/computed tomography, performed at six months post-treatment, revealed a good local response and no distant metastases. The patient was placed on routine follow up at two-monthly intervals. Two months into follow up, he presented with a right-sided oculomotor nerve palsy and partial Horner's syndrome. Imaging and biopsy revealed a pituitary fossa metastasis (p16-negative squamous cell carcinoma), and a further positron emission tomography/computed tomography visualised this lesion. He was deemed unsuitable for further intervention and underwent palliative radiotherapy for symptom control. CONCLUSION: This case represents the first reported isolated pituitary fossa metastasis from a tonsillar squamous cell carcinoma. A high degree of clinical suspicion is recommended, along with a low threshold for biopsy and a cautioned use of positron emission tomography/computed tomography, when investigating such patients.

Exploration of a New Source of Sustainable Nanomaterial from the Koh-e-Suleiman Mountain Range of Pakistan for Industrial Applications.

The present study aimed to explore a new source of montmorillonite and to develop an extraction and purification protocol for its isolation from raw clay samples acquired from the Koh-e-Suleiman mountain range in Pakistan. The process involved the collection of raw clay from the source, identification and quantification of montmorillonite. Granulometric extraction and purification protocols increased the montmorillonite content from 21.8-25.1% in the raw clay to 90.1-93.9% after small-scale extraction and 85.33-89.33% on a larger scale. A techno-economic analysis highlighted the practicality and economic benefits of large-scale extraction for industrial applications. This study highlights the existence of a substantial new source of this valuable clay which is currently used across multiple industries including construction, pottery making, pharmaceuticals, cosmetics and engineering. It is intuitively expected that the large-scale extraction of the material will improve the economic condition of the region by providing employment opportunities to locals and may be a valuable resource for export.

Lipid droplet analysis using in vitro bovine oocytes and embryos.

The aim of this study was to quantify the content of lipid droplets in bovine oocytes and embryos from Bos indicus (Bi), Bos taurus (Bt) and Bos indicus × Bos taurus (Bi × Bt). Oocytes were aspirated post-mortem and subjected to in vitro maturation, in vitro fertilization and in vitro development; the medium employed at each stage (TCM-199, TALP, SOF) was supplemented with (i) serum replacement (SR), (ii) foetal calf serum (FCS) or (iii) oestrous cow serum (ECS). The structure and distribution of the lipid droplets were established using electron microscopy, but were quantified using an optical microscope on semi-fine toluidine blue-stained sections. The highest percentage of embryos corresponded to those produced with FCS and ECS, which differed from embryos generated with SR (p < 0.05). The highest percentage of morulae and the lowest percentage of blastocysts were obtained with the SR supplement (p < 0.05). The oocytes cultured in FCS demonstrated a higher number of lipid droplets compared to those cultured in SR and ECS (p < 0.05). Less accumulation of lipids was observed in embryos supplemented with SR. The lowest and highest numbers of lipid droplets in oocytes corresponded to the Bi and Bt strain, respectively. The lowest amount of lipid droplets in embryos was observed in Bi (p < 0.05). In conclusion, supplementation of the in vitro development culture medium (synthetic oviduct fluid) with a synthetic substitute serum produced similar results in terms of embryo development compared to those obtained with FCS, but a decreased degree of lipid droplet accumulation was observed in the in vitro-cultured embryos.

Seasonal effect on zebu embryo quality as determined by their degree of apoptosis and resistance to cryopreservation.

In order to optimize the production of embryos under tropical conditions and to test a possible seasonal effect on embryo quality, 40 Zebu cows were superovulated during the dry season (April to May) and during the rainy season (July to August). A total of 116 (average 2.7/cow) and 83 embryos (3.5 average/cow) were obtained during the respective seasons. After classification as good, fair or poor quality, embryos were tested based on their ultrastructural differences (n = 53 dry season 16 good, 20 fair and 17 poor and n = 61 rainy season 21 good, 20 fair and 20 poor) and their degree of apoptosis using the TUNEL technique (n = 30 during the dry season and n = 55 in the rainy season). Structural characteristics determining embryo quality varied between good and fair quality embryos. No difference, however, was observed between good, fair and poor quality embryos from the two seasons. The number of TUNEL-positive cells was different among embryos (p < 0.001), being lower in labelled cells of good quality embryos regardless of the season. Fewer apoptotic cells were observed in embryos assigned in all three quality levels during the rainy season (p < 0.001). Ultrastructural evaluations confirmed the results obtained by TUNEL. Cryopreserved embryos of good (n = 25 in each season) and fair quality (n = 11 dry season; n = 17 rainy season) showed a significant decrease of TUNEL-positive cells during the rainy season (p < 0.05). Results suggest that embryos collected in the dry season have more cellular damage in contrast; embryos cryopreserved in the rainy season appeared morphologically better equipped to result in a pregnancy following transfer.

Conduction failure of action potentials in sensory sural nerves of undernourished rats.

In order to determine possible functional and morphometrical alterations produced by perinatal undernourishment on peripheral nerves, sensory sural nerves from control and undernourished rats of 30 and 90 postnatal days of age were dissected and divided in two segments, one for recording the compound action potential (CAP) and the other for histological examination. Nerves from undernourished animals showed maximal CAP responses of smaller amplitude and area, larger trial-to-trial variability in area, and a significant reduction in axonal myelin sheath thickness than nerves from control animals. It is suggested that perinatal undernourishment produces changes in axonal myelin sheath structure, resulting in severe alterations in the generation and propagation of action potentials (block and/or intermittent conduction) in sensory afferent fibers in the rat.

Neurophysiology of the parieto-frontal system during target interception.

We studied the functional properties of neurons of two elements of the parieto-frontal system: area 7a of the PPC and the motor cortex (M1), during an interception task of stimuli moving in real (RM) and apparent motion (AM). The stimulus moved along a circular path with one of 5 speeds, and was intercepted at 6 o'clock by exerting a force pulse on a joystick. A smooth stimulus motion was produced in RM, whereas in AM 5 stimuli were flashed successively at the vertices of a pentagon. The results showed, that a group of neurons in both areas above responded not only during the interception but also during a NOGO task in which the same stimuli were presented in the absence of a motor response. Most of these neurons were tuned to the stimulus angular position. In addition, we found that the time-varying neuronal activity in both areas was related to various aspects of stimulus motion and hand force, with stimulus-related activity prevailing in area 7a and hand-related activity prevailing in M1. Interestingly, the neural activity was selectively associated with the stimulus angle during RM, whereas it was tightly correlated to the time-to-contact during AM. Thus, the results suggest that area 7a was processing high level features of the circularly moving stimuli and was involved in the production of an early command signal for stimulus interception, whereas M1 was still processing some aspect of the visual stimulus that were used to trigger the interception movement using a predictive mechanism.

Comparison of stereoscopy, light microscopy and ultrastructural methods for evaluation of bovine embryos.

The most important point in embryo transfer success is the evaluation of the stage of development and quality of embryos. Therefore, the purpose of this study was to compare the morphological evaluation of embryos using stereoscopy, light microscopy and electron microscopy in order to establish the accuracy of former method compared with more invasive and accurate procedures. For this purpose, 23 Brahman x Swiss cows were used and synchronized with Norgestomet 6 mg plus, 5 mg Estradiol valerate (Syncromate B(R), Rhone Merieux, Mexico, Mexico City) and superovulated with Folltropin-V 240 mg (Vetrepharm, Mexico, Mexico City). Non-surgical embryo collection was performed 7.5 days after insemination. Descriptive statistics analysis was used to assess the data. Seventy-eight embryos were collected and classified by stereoscopic microscopy, finding 51.2% (40) of good quality, 25.6% (20) fair and 24.3% (19) poor. Later, under light microscopy observation, evaluation of the same embryos resulted in 25.6% (20) good, 32.0% (25) fair and 42.3% poor quality. Finally, in the evaluation of embryos under electron microscopy 24.3% (19) were found to be of good quality, 29.3% (23) fair and 46.1% (36) poor. Evaluation of embryos with stereoscopic microscopy was found to be very subjective, as nearly 50% of embryos classified by this method as good quality, showed features of degenerative stages under light and electron microscopy. Embryos with these features are generally frozen and transferred, which could be one of the reasons for having low fertility rate in embryo transfer programmes.

Assessment of the in vitro and in vivo biological activities of the human follicle-stimulating isohormones.

Gonadotropins are synthesized and released in different molecular forms. In this article, we present evidence that the glycosylation variants of human pituitary FSH exhibit differential and divergent effects at the target cell level and that less sialylated, short-lived variants may exert significant effects in in vivo conditions. Less acidic/sialylated glycoforms (elution pH value 6.60-4.60 as disclosed by high resolution chromatofocusing of anterior glycoprotein extracts), induced higher cAMP release, estrogen production and tissue-type plasminogen activator (tPA) enzyme activity as well as cytochrome P450 aromatase and tPA mRNA expression in cultured rat granulosa cells than the more acidic analogs (pH<4.76). By contrast, the more acidic/sialylated glycoforms induced higher alpha-inhibin subunit mRNA expression than their less acidic counterparts. In cumulus enclosed oocytes isolated from mice ovaries, addition of less acidic isoforms induced resumption of meiosis more efficiently than the more acidic analogs. Interestingly, the least acidic isoform (pH>7.10) behave as a strong antagonist of several FSH-mediated effects. Assessment of the in vivo effects of the isoforms on granulosa cell proliferation in follicles from immature rats, revealed that short-lived isoforms were equally or even more efficient than their more acidic counterparts in maintaining granulosa cell proliferation when administered immediately after hypophysectomy. These results show that the naturally occurring human FSH isoforms may exhibit differential or even unique effects at the target cell level and that factors other than the metabolic clearance rate of the molecule (including receptor-binding affinity and capability of the ligand to activate its receptor and trigger intracellular signaling) also play an important role in determining the net in vivo effects of a particular FSH variant.

Effects of optic flow in motor cortex and area 7a.

Moving visual stimuli were presented to behaving monkeys who fixated their eyes and did not move their arm. The stimuli consisted of random dots moving coherently in eight different kinds of motion (right, left, up, downward, expansion, contraction, clockwise, and counterclockwise) and were presented in 25 square patches on a liquid crystal display projection screen. Neuronal activity in the arm area of the motor cortex and area 7a was significantly influenced by the visual stimulation, as assessed using an ANOVA. The percentage of cells with a statistically significant effect of visual stimulation was 3 times greater in area 7a (370/587, 63%) than in motor cortex (148/693, 21.4%). With respect to stimulus properties, its location and kind of motion had differential effects on cell activity in the two areas. Specifically, the percentage of cells with a significant stimulus location effect was approximately 2.5 times higher in area 7a (311/370, 84%) than in motor cortex (48/148, 32.4%), whereas the percentage of cells with a significant stimulus motion effect was approximately 2 times higher in the motor cortex (79/148, 53.4%) than in area 7a (102/370, 27.6%). We also assessed the selectivity of responses to particular stimulus motions using a Poisson train analysis and determined the percentage of cells that showed activation in only one stimulus condition. This percentage was 2 times higher in the motor cortex (73.7%) than in area 7a (37.7%). Of all kinds of stimulus motion tested, responses to expanding optic flow were the strongest in both cortical areas. Finally, we compared the activation of motor cortical cells during visual stimulation to that observed during force exertion in a center --> out task. Of 514 cells analyzed for both the motor and visual tasks, 388 (75.5%) showed a significant relation to either or both tasks, as follows: 284/388 (73.2%) cells showed a significant relation only to the motor task, 27/388 (7%) cells showed a significant relation only to the visual task, whereas the remaining 77/388 (19.8%) cells showed significant relations to both tasks. Therefore a total of 361/514 (70.2%) cells were related to the motor task and 104/514 (20.2%) were related to the visual task. Finally, with respect to receptive fields (RFs), there was no clear visual receptive field structure in the motor cortical neuronal responses, in contrast to area 7a where RFs were present and could be modulated by the type of optic flow stimulus.

Comparative characterization of cell death between Sf9 insect cells and hybridoma cultures.

Physiological cell death (PCD) in Sf9 insect cell batch cultures was comprehensively characterized using simultaneous determinations of qualitative and quantitative assays, including agarose gel electrophoresis, confocal, epifluorescence, and transmission electron microscopy, and DNA content by flow cytometry. Results were compared to hybridoma cultures where abundant information of apoptosis exists. Both cultures shared some typical apoptosis features, including cell shrinkage, loss of sphericity, swollen endoplasmic reticulum and Golgi apparatus, chromatin condensation, and specific DNA degradation. However, distinctive morphological and kinetic differences between both cultures revealed that Sf9 cells died by an atypical PCD process characterized by absence of nuclear fragmentation, scarce association of condensed chromatin to the nuclear envelope, swollen mitochondria, and high nonspecific DNA degradation. These features, distinctive of necrosis, were not observed in the normal apoptotic process of hybridomas. Glucose depletion marked the appearance of apoptotic Sf9 cells, which there up on increased gradually, whereas apoptotic hybridomas rapidly increased upon glutamine depletion. Furthermore, active phagocytosis was found in Sf9 viable cells, a characteristic phenomenon during in vivo apoptosis but uncommon for in vitro cultures. Sf9 cells contained unusually high numbers of phagosomes, particularly after glucose depletion. Additionally, few apoptotic bodies accumulated in culture, suggesting their elimination by phagocytosis. Other distinctive characteristics of Sf9 cells were the presence of a polynucleated hypertrophic population fraction, polyploidy, cell cycle arrest in G2/M phase, and more necrosis compared to hybridomas. Such phenomena prevented a reliable quantification of apoptosis from determination of the sub-G1 peak. Nonetheless, emergence of a bimodal Sf9 cell size distribution coincided with the increase in the sub-G1 population and onset of death. The fraction of particles in the smaller peak (6-11 microm diameter) closely correlated with the fractions of apoptotic bodies, late apoptotic, and secondary necrotic cells. Accordingly, Sf9 cell size was shown to be an effective, rapid, and simple parameter for quantifying death. Altogether, the results of this study provide new insights into PCD and other phenomena in insect cell culture important for biotechnological applications of Sf9 cells.

Effect of perinatal food deficiencies on the compound action potential evoked in sensory nerves of developing rats.

The aim of this study was to analyze the possible alterations produced by inadequate perinatal food intake, in quantity (undernutrition) or quality (malnutrition), on the generation and propagation of the compound action potential (CAP) evoked in sensory sural nerves, during the postnatal development of the rat. Low intensity stimulation (2-3 times the threshold of the most excitable nerve fibers; xT) of the sural nerve evoked an early potential (CAP-A component) which is due to activation of low-threshold, fast-conducting myelinated group A afferent fibers. Meanwhile, at higher stimulus intensity (20-30T) it produced a second, long-lasting potential (CAP-C component) probably due to activation of high-threshold, slow-conducting group Adelta or C afferent fibers. Compared to control nerves, the CAP-A component, but not the CAP-C component of undernourished and malnourished nerves showed significant changes in amplitude, area, electrical threshold and conduction velocity (except absolute refractory period) at several postnatal ages. Our results may suggest that a relative large number of myelinated group A afferent fibers in the sural nerve of undernourished and malnourished animals suffer severe alterations on their electrophysiological properties of generation and propagation of the action potential during the postnatal development of the rat.

A recombinant vaccinia virus containing the papilloma E2 protein promotes tumor regression by stimulating macrophage antibody-dependent cytotoxicity.

Human papillomavirus infection is associated with cervical cancer. The E6 and E7 papillomavirus proteins are normally required for the maintenance of the malignant phenotype. Expression of these proteins in infected cells is negatively regulated by the binding of the papilloma E2 protein to the long terminal control region of the papilloma virus genome. The E2 protein can also promote cell arrest and apoptosis in HeLa cells. Therefore, it is clear that this protein has the potential of inhibiting the malignant phenotype. Because, anticancer vaccines based in vaccinia viruses have recently been shown to be an effective way to treat and to eradicate tumors, a recombinant vaccinia virus expressing the E2 gene of bovine papilloma virus (Modified Vaccinia Ankara, MVA E2) was created, to explore further the antitumor potential of the E2 protein. A series of rabbits, containing the VX2 transplantable papilloma carcinoma, were treated with MVA E2. An impressive tumor regression, up to a complete disappearance of tumor, was observed in most animals (80%). In contrast, very little or no regression was detected if the normal vaccinia virus was used. Lymphocytes isolated from MVA E2-treated rabbits did not show cytotoxic activity against tumor cells. However, in these animals a humoral immune response against tumor cells was observed. These antitumor antibodies were capable of activating macrophages to destroy tumor cells efficiently. These data indicate that injecting the MVA E2 recombinant vaccinia virus directly into the tumor results in a robust and long-lasting tumor regression. Data also suggest that antitumor antibodies are responsible, at least in part, for eliminating tumors by activating macrophage antibody-dependent cytotoxicity.

Human tumor growth is inhibited by a vaccinia virus carrying the E2 gene of bovine papillomavirus.

BACKGROUND: Papillomavirus is the etiologic agent associated with cervical carcinoma. The papilloma E2 protein is able to regulate negatively the expression of E6 and E7 papilloma oncoproteins. Therefore, a new, highly attenuated vaccinia virus known as modified vaccinia virus Ankara (MVA), which carries the papillomavirus E2 gene, was used for the treatment of tumors associated with human papillomavirus. METHODS: Analysis of expression of the E2 gene from the recombinant vaccinia virus was performed by reverse transcription-polymerase chain reaction of RNA isolated from infected cells. Detection of the E2 protein was done by immunoprecipitation from proteins labeled with [(35)S]-methionine, isolated from infected cells. The therapeutic effect of the MVA E2 recombinant virus over human tumors was tested in nude mice bearing tumors generated by inoculation of HeLa cells. Series of 10 nude mice with tumors of different sizes were injected with MVA, MVA E2, or phosphate-buffered saline. Tumor size was monitored every week to assess growth. RESULTS: The MVA E2 recombinant virus efficiently expressed the E2 protein in BS-C-1 cells. This protein was able to repress, in vivo, the papillomavirus P105 promoter, which controls the expression of the E6 and E7 oncoproteins. In nude mice the MVA E2 virus reduced tumor growth very efficiently. In contrast, tumors continued to grow in mice treated with MVA or PBS. The life expectancy of MVA E2-treated mice was also increased three- to fourfold compared with that of animals that received MVA or PBS. CONCLUSIONS: The growth of human tumors was efficiently inhibited by the MVA E2 recombinant vaccinia virus. The absence of side effects in treated animals suggested that the MVA E2 virus is a safe biologic agent that could in the future be used in humans for the treatment of cervical carcinoma.

Bilioma secondary to choledocholithiasis.

Bilioma secondary to choledocholithiasis is rare. We report a patient in whom a large common bile duct stone was responsible for leak from the infraduodenal segment of the bile duct. Choledochotomy with extraction of stone followed by T-tube drainage of the bile duct and evacuation of the bilioma resulted in complete recovery.

Categorical perception of somesthetic stimuli: psychophysical measurements correlated with neuronal events in primate medial premotor cortex.

In this paper we describe a type of neuron of the medial premotor cortex (MPC) that discharged differentially during a categorization task and reflected in their activity whether the speed of a tactile stimulus was low or high. The activity of these neurons was recorded in the MPC contralateral (right MPC, n = 88) and ipsilateral (left MPC, n = 103) to the stimulated hand of four monkeys performing this somesthetic task. Animals performed the task by pressing with the right hand one of two target switches to indicate whether the speed of probe movement across the skin of the left hand was low or high. Differential responses of MPC neurons occurred during the stimulus and reaction time period. We used an analysis based on signal detection theory to determine whether these differential responses were associated with the animal's decision. According to this analysis, 104 of the 191 neurons (right MPC, n = 48; left MPC, n = 56) coded the categorization of the stimulus speeds (categorical neurons). In a light instruction task, we tested the possibility that the categorical neurons (n = 71) were associated with the intention to press, or with the trajectory of the hand to one of the two target switches used to indicate categorization. In this situation, each trial began as in the somesthetic categorization task, but one of the two target switches was illuminated beginning with the skin indentation, continued during the delay period and turned off when the probe was lifted off from the skin. This condition instructed the animal which target switch was required to be pressed for reward. Very few neurons (14 of 71) maintained their differential responses observed in the categorization task. Some categorical neurons (n = 5) were also studied; the animal categorized the tactile stimulus speeds, but knew in advance whether the stimulus speed was low or high (categorization + light instruction). This was made by illuminating one of the two target switches which was associated with the stimulus speed. The categorical response was considerably attenuated in this condition. Interestingly, during the delay period, these neurons reflected in their activity whether the stimulus was low or high. A number of the categorical MPC neurons (n = 30) were studied when the same set of stimuli, used in the categorization, were delivered passively. None of these neurons responded in this condition. These results suggest that the MPC, apart from its well-known role in motor behavior, is also involved in the animal's decision during the execution of this learned somesthetic task.

Role of primary somatic sensory cortex in the categorization of tactile stimuli: effects of lesions.

We lesioned the right primary somatic sensory (S1) cortex in two monkeys trained to categorize the speed of moving tactile stimuli. Animals performed the task by pressing with the right hand one of two target switches to indicate whether the speed of a probe moving across the glabrous skin of the left hand was low or high. Sensory performance was evaluated with psychometric techniques and motor behavior was monitored by measuring the reaction (RT) and movement (MT) times before the experiment and throughout the 60 days after the ablation of SI cortex. After the lesion, there was a slight increase in the RTs but no change in the MTs, indicating that removal of SI cortex did not affect the animals' capacity to detect the stimuli. However, monkeys lost their ability to categorize the stimulus speeds. This effect was observed from the 1st day after the lesion until the end of the study. We conclude that somatosensory areas outside SI can by themselves process tactile information in a limited way and that the extraction of higher-order features that takes place during the categorization task requires the intervention of SI cortex.