Fine needle aspiration cytology of hepatic metastasis from a meningeal hemangiopericytoma. A case report

BACKGROUND: Hemangiopericytomas (HPCs) are rare spindle cell tumors, constituting 2.5% of soft tissue neoplasms. Few reports have addressed the fine needle aspiration (FNA) cytology of HPC. CASE: We describe the FNA biopsy (FNAB) findings in a 44-year-old patient with a previously resected meningeal hemangiopericytoma. The patient underwent ultrasound-guided FNAB of a 16.0-cm, radiographically heterogeneous density in the liver. The FNA smear showed crowded, ovoid to spindle-shaped cells with poorly defined, scant cytoplasm. The neoplastic cells were positive for CD34 and negative for CD31, factor VIII, glial fibrillary acid protein and cytokeratin AE1/AE3, supporting a diagnosis of HPC and compatible with metastasis from the patient's cerebral tumor. CONCLUSION: This case documents the role of FNA cytology in confirming HPC.     

High-precision,high-throughput stability determinations facilitated by robotics and a semiautomated titrating fluorometer

The use of statistical modeling to test hypotheses concerning the determinants of protein structure requires stability data (e.g., the free energy of denaturation in H(2)O, DeltaG(HOH)) from hundreds of protein mutants. Fluorescence-monitored chemical denaturation provides a convenient method for high-precision, high-throughput DeltaG(HOH) determination. For eglin c we find that a throughput of about 20 min per protein can be attained in a two-channel semiautomated titrating fluorometer. We find also that the use of robotics for protein purification and preparation of the solutions for chemical denaturation gives highly precise DeltaG(HOH) values in which the standard deviation of values from multiple preparations (+/-0.051 kcal/mol) differs very little from multiple measurements from a single preparation (+/-0.040 kcal/mol). Since the variance introduced into model fitting by DeltaG(HOH) increases as the square of measurement error, there is a premium on precision. In fact, the fraction of stability behavior explicable by otherwise perfect models goes from 98% to only 50% over the error range commonly reported for chemical denaturation measurements (0.1-0.6 kcal/mol). We have found that the precision of chemical denaturation DeltaG(HOH) measurements depends most heavily on the precision of the instrument used, followed by protein purity and the capacity to precisely prepare the solutions used for titrations.     

Dynamin2 and cortactin regulate actin assembly and filament organization

The GTPase dynamin is required for endocytic vesicle formation. Dynamin has also been implicated in regulating the actin cytoskeleton, but the mechanism by which it does so is unclear. Through interactions via its proline-rich domain (PRD), dynamin binds several proteins, including cortactin, profilin, syndapin, and murine Abp1, that regulate the actin cytoskeleton. We investigated the interaction of dynamin2 and cortactin in regulating actin assembly in vivo and in vitro. When expressed in cultured cells, a dynamin2 mutant with decreased affinity for GTP decreased actin dynamics within the cortical actin network. Expressed mutants of cortactin that have decreased binding of Arp2/3 complex or dynamin2 also decreased actin dynamics. Dynamin2 influenced actin nucleation by purified Arp2/3 complex and cortactin in vitro in a biphasic manner. Low concentrations of dynamin2 enhanced actin nucleation by Arp2/3 complex and cortactin, and high concentrations were inhibitory. Dynamin2 promoted the association of actin filaments nucleated by Arp2/3 complex and cortactin with phosphatidylinositol 4,5-bisphosphate (PIP2)-containing lipid vesicles. GTP hydrolysis altered the organization of the filaments and the lipid vesicles. We conclude that dynamin2, through an interaction with cortactin, regulates actin assembly and actin filament organization at membranes.     

Interaction with GM130 during HERG ion channel trafficking. Disruption by type 2 congenital long QT syndrome mutations. Human Ether-à-go-go-Related Gene

Many mutations in the Human Ether-à-go-go-Related Gene (HERG) cause type 2 congenital long QT syndrome (LQT2) by disrupting trafficking of the HERG-encoded potassium channel. Beyond observations that some mutations trap channels in the endoplasmic reticulum, little is known about how trafficking fails. Even less is known about what checkpoints are encountered in normal trafficking. To identify protein partners encountered as HERG channels are transported among subcellular compartments, we screened a human heart library with the C terminus of HERG using yeast two-hybrid technology. Among the proteins isolated was GM130, a Golgi-associated protein involved in vesicular transport. The interaction mapped to two non-contiguous regions of HERG and to a region just upstream of the GRASP-65 interaction domain of GM130. GM130 did not interact with the N or C terminus of either KvLQT1 or Shaker channels. LQT2-causing mutations in the HERG C terminus selectively disrupted interactions with GM130 but not Tara, another HERG-interacting protein. Native GM130 and stably expressed HERG were co-immunoprecipitated from HEK-293 cells using GM130 antibodies. In rat cardiac myocytes and HEK-293 cells, confocal immunocytochemistry showed co-localization of GM130 and HERG to the Golgi apparatus. Overexpression of GM130 suppressed HERG current amplitude in Xenopus oocytes, as if by providing an excess of substrate at the Golgi checkpoint. These findings indicate that GM130 plays a previously undefined role in cargo transport. We propose that the cytoplasmic C terminus of HERG participates in the tethering or possibly targeting of HERG-containing vesicles within the Golgi via its interaction with GM130.     

Telomere length deregulation and enhanced sensitivity to genotoxic stress in Arabidopsis mutants deficient in Ku70

The Ku70/80 heterodimer is a critical component of the non-homologous end-joining (NHEJ) pathway and of the telomere cap in yeast and mammals. We report the molecular characterization of the KU70 and KU80 genes in Arabidopsis and describe the consequences of a Ku70 deficiency. Arabidopsis KU70/80 genes are ubiquitously expressed and their products form stable heterodimers in vitro. Plants harboring a T-DNA insertion in KU70 exhibit no growth or developmental defects under standard growth conditions. However, mutant seedlings are hypersensitive to gamma-irradiation-induced double-strand breaks. Unexpectedly, we found that mutants are hypersensitive to methyl methanosulfonate during seed germination, but lose this sensitivity in seedlings, implying that the requirement for NHEJ varies during plant development. Lack of Ku70 results in a dramatic deregulation of telomere length control, with mutant telomeres expanding to more than twice the size of wild type by the second generation. Furthermore, in contrast to the situation in mammals, chromosome fusions are not associated with a Ku deficiency in Arabidopsis. These findings imply that Ku may play a different role in capping plant and animal telomeres.     

Angiotensin-(1-7) and bradykinin interaction in diabetes mellitus: in vivo study

The interaction between angiotensin-(1-7) (Ang-(1-7)) and bradykinin (BK) was determined in the mesentery of anesthetized Wistar alloxan-diabetic and non-diabetic rats using intravital microscopy. Impaired BK vasodilation observed in arterioles of diabetic rats was restored by acute and chronic insulin treatment as well as by enalapril. Though capable of potentiating BK in non-diabetic rats, Ang-(1-7) did not potentiate BK in diabetic rats. Chronic but not acute insulin treatment restored the potentiation, whereas enalapril did not. Potassium channel blockade (by tetraethylammonium (TEA)) but not nitric oxide (NO) synthase inhibition (by N-omega-nitro-L-arginine-methyl-esther (L-NAME)) abolished the restorative effect of insulin. Our data allow us to suggest that the alteration observed is restored by insulin by a mechanism involving membrane hyperpolarization but not NO release. The beneficial effect of enalapril in diabetes might not involve the potentiation of BK by Ang-(1-7).     

Spontaneous gallstone formation in deer mice: interaction of cholesterol,bile acids,and dietary fiber

A study of the physiologic and ecologic factors involved in a spontaneous seasonal gallstone cycle of deer mice (Peromyscus maniculatus gambelii) was conducted at the Tulelake National Wildlife Refuge (California, USA) from March 1991 to June 1992. The specific hypothesis examined was whether or not seasonal increases in dietary fiber intake provides the necessary conditions for a solubility defect, or supersaturation mechanism, resulting in precipitation of cholesterol gallstones. Results indicated that in addition to the seasonal gallstone prevalence cycle, these deer mice exhibit significant seasonal cycling in serum cholesterol, serum bile acids, fecal bile acids, and diet composition. These physiologic and dietary cycles were phase-advanced 3 mo over the gallstone prevalence cycle, indicating an approximate 3 mo time period for gallstone formation under field conditions. Further, seasonal dietary fiber (plant material and seeds) was positively correlated with both serum cholesterol and the fecal bile acids. This suggests that in wild deer mice, variations in dietary fiber may significantly affect the resorption of bile acids, thereby providing a potential physiologic and nutritional mechanism for spontaneous cholesterol gallstone formation.     

Role of mass spectrometry in the purification of peptides and proteins

Experiments were carried out to evaluate the fractionation of proteins and peptides according to mass. Model mixtures were separated by either reversed-phase or ion-exchange chromatography with mass spectrometry-compatible mobile phase additives. Fraction collection was triggered by the mass/charge ratio of each one of the components of the mixture. Chromatography was additionally monitored with a UV-Vis detector in order to compare the new technique with generally accepted in separations. The results indicated that adequate purification is achieved by this new technique. Fraction collection triggered by changes in the mass/charge ratio reduces sample handling and analysis time. This study demonstrates the utility of mass-directed fractionation of peptides and proteins when mass spectrometry-compatible mobile phase additives are used.