Segregation of all four major fibrillar collagen genes in the Marfan syndrome.

Linkage markers at or close to the genes encoding the three major fibrillar collagens were used to analyze the segregation of these loci in six pedigrees with dominantly inherited Marfan syndrome. Four pedigrees were discordant at one of the Type I collagen loci (COL1A2), and, of these, two were discordant at the other Type I locus (COL1A1). The Marfan syndrome also segregated independently of the structural loci for Type II and Type III collagen in these two families. This is evidence against the Marfan syndrome being, in general, due to mutations in the major fibrillar collagen genes.     

In situ hybridization using 32P labelled oligodeoxyribonucleotides for the cellular localisation of mRNA in neuronal and endocrine tissue

Summary Methodological variables for in situ hybridization using ³²P labelled oligodeoxyribonucleotides (oligomers) have been examined. Four different oligomers directed against proglucagon messenger RNA (mRNA) and two different oligomers against prosomatostatin mRNA have been used. Specific hybridization was obtained in adult rat brain, stomach and pancreas and in neonatal rat ileum. Tissue was perfusion fixed with 4% paraformaldehyde 0.2% glutaraldehyde and hybridization was carried out in 50% formamide for 72 h at 42° C. Using hybridization conditions of lower stringency (33% formamide) labelling was also obtained in guinea pig tissue. Other variables which affected hybridization signal intensity were the inclusion of a prehybridization dehydration stage, the probe concentration, the inclusion of ammonium acetate in the posthybridization dehydrating ethanols and in the autoradiographic emulsion, and the exposure time. The localisation of proglucagon mRNA in rat pancreas using a 20mer was used as a model tissue for testing these methodological variables and the results were found generally also to apply to the other probes and tissues tested. The methods described provide single cell resolution and show that ³²P labelled oligomers may be used to localise neuropeptide and endocrine mRNAs in different types of tissue and in different mammalian species.     

Lysophosphatidylcholine uptake and metabolism in the adult rabbit lung

Tracer quantities of 3H-labeled lysoPC and 32P-labeled natural rabbit surfactant were given intratracheally via a bronchoscope and [14C]palmitate was given intravenously to 25 rabbits with labeled PC and lysoPC measured in the alveolar wash, lung homogenate, lamellar bodies and microsomes at five times from 10 min to 6 h after tracheal injection. Surprisingly, only 31% of the administered lysoPC remained in its original form in the total lungs (alveolar wash + lung homogenate) by 10 min, of which 77% was in the alveolar wash. Meanwhile, by 10 min an additional 37% was already converted to PC, of which more than 98% was in the lung homogenate. LysoPC continued to be rapidly and efficiently converted to PC, with 62% conversion measured at 3 h. The converted lysoPC initially appeared with high specific activity in microsomes, then in lamellar bodies, and finally in the alveolar wash. The intravascular palmitate labeled lung PC had similar specific activity-time profiles in the subcellular fractions, while intratracheally administered natural rabbit surfactant had a constantly low specific activity in microsomes and much higher specific activities in lamellar bodies and alveolar wash. Another 25 rabbits received intratracheal lysoPC labeled in both the choline and palmitate moieties and then were studied from 1 to 24 h after tracheal injection. The ratio of the palmitate to choline labels indicated uptake and conversion to PC primarily by direct acylation rather than transacylation and by intact reuptake and conversion rather than breakdown and resynthesis. LysoPC is an attractive 'metabolic probe' of surfactant metabolism which undergoes very rapid and efficient intracellular conversion to PC via a subcellular pathway that parallels the remodeling and de novo synthetic pathways.     

Organic free radical levels in seeds and pollen: The effects of hydration and aging

In view of their possible role in oxidative deterioration of seeds and pollen, organic free radicals were measured by electron spin resonance in embryonic axes and cotyledons of soybean [ Glycine max (L.) Merr], embryo and endosperm fractions of corn [ Zea mays L.] and pollen of cattail [ Typha latifolia L.]. A pronounced decline in the radical signal ensued when hydration increased above about 7% (wet weight basis) in both the seed materials and in pollen. Moderate hydration of the soybean axis followed by drying led to a small decrease in organic free radicals compared to untreated material, especially if the desiccation step was performed under nitrogen. In a comparison of soybeans of various ages under normal storage, organic free radical levels in the axis showed little or no increase with age. In marked contrast, over 5 days of accelerated aging at 40°C and near-saturating humidity, organic radical levels approximately doubled in the axis. This pronounced increase in free radical content was not associated with a decrease in the proportion of polyunsaturated fatty acids. The data suggest that hydration of seed and pollen causes a release of free radicals from the trapped state.     

Surface charge on isolated maize-coleoptile amyloplasts

Whole amyloplasts were isolated from Zea mays L. coleoptiles. Electron microscopy confirmed that their envelopes consisted of two intact membranes. Their surface charge was quantified through electrokinetic measurements employing a vertically oriented cataphoresis cell. Amyloplasts from coleoptiles of different ages (5–9 d) and degree of exposure to light (0–9 d) all had negative zeta potentials (mean of -19.4 mV). Isolated starch granules had comparable values. The net negative surface charge was confirmed ultrastructurally by the binding of cationised ferritin to both amyloplasts and starch grains. Cationised ferritin tagged with a fluorescent label (fluorescein isothiocyanate) showed binding to some amyloplasts but not to starch grains. These results are discussed in the context of a hypothesized role for statolith charge in plant graviperception.     

Synaptic connections of substance P-immunoreactive nerve terminals in the substantia nigra of the rat

Summary A monoclonal antibody that recognises the C-terminal part of substance P was used to study immunoreactive structures in the substantia nigra by the unlabeled antibody, peroxidase-antiperoxidase procedure. Immunoreactivity was present in nerve fibres in all parts of the substantia nigra, particularly in the pars reticulata and pars lateralis. Electron microscopically two types of bouton immunoreactive for substance P were found: Type 1 contained large electron-lucent vesicles, occasional large granulated vesicles and formed symmetrical synapses with dendrites. Type 2 boutons contained smaller, round electron-lucent vesicles, many large granular vesicles and formed asymmetrical synapses (having prominent postjunctional dense bodies) with dendrites and perikarya.Immunoreactive fibres with varicosities that had been identified light microscopically were studied in serial sections in the electron microscope. Each identified varicosity contained synaptic vesicles and formed a single synapse. An individual fibre formed boutons of only one kind (type 1 or type 2) and could form multiple synapses with the same neuron. Thus, an identified fibre in the pars compacta had eight varicosities, each of which was in synaptic contacts (type 2) with the dendrites or soma of the same neuron.The results are consistent with the concept that substance P is a synaptic transmitter in the substantia nigra and indicate that neurons in this region may receive a significant input from substance P-containing afferents, and that there are at least two types of such afferent fibres.     

Biodeterioration of Mayan buildings at uxmal and tulum,Mexico

Uxmal and Tulum are two important Mayan sites in the Yucatan peninsula. The buildings are mainly composed of limestone and grey/black discoloration is seen on exposed walls and copious greenish biofilms on inner walls. The principal microorganisms detected on interior walls at both Uxmal and Tulum were cyanobacteria; heterotrophic bacteria and filamentous fungi were also present. A dark‐pigmented mitosporic fungus and Bacillus cereus, both isolated from Uxmal, were shown to be acidogenic in laboratory cultures. Cyanobacteria belonging to rock‐degrading genera Synechocystis and Gloeocapsa were identified at both sites. Surface analysis previously showed that calcium ions were present in the biofilms on buildings at Uxmal and Tulum, suggesting the deposition of biosolubilized stone. Apart from their potential to degrade the substrate, the coccoid cyanobacteria supply organic nutrients for bacteria and fungi, which can produce organic acids, further increasing stone degradation.     

Nonbenzamidine acylsulfonamide tissue factor–factor VIIa inhibitors

Aminoisoquinoline and isoquinoline groups have successfully replaced the more basic P1 benzamidine group of an acylsulfonamide factor VIIa inhibitor. Inhibitory activity was optimized by the identification of additional hydrophobic and hydrophilic P′ binding interactions. The molecular details of these interactions were elucidated by X-ray crystallography and molecular modeling. We also show that decreasing the basicity of the P1 group results in improved oral bioavailability in this chemotype.     

Multimodal pressure-flow method to assess dynamics of cerebral autoregulation in stroke and hypertension

Background

This study evaluated the effects of stroke on regulation of cerebral blood flow in response to fluctuations in systemic blood pressure (BP). The autoregulatory dynamics are difficult to assess because of the nonstationarity and nonlinearity of the component signals.

Methods

We studied 15 normotensive, 20 hypertensive and 15 minor stroke subjects (48.0 ± 1.3 years). BP and blood flow velocities (BFV) from middle cerebral arteries (MCA) were measured during the Valsalva maneuver (VM) using transcranial Doppler ultrasound.

Results

A new technique, multimodal pressure-flow analysis (MMPF), was implemented to analyze these short, nonstationary signals. MMPF analysis decomposes complex BP and BFV signals into multiple empirical modes, representing their instantaneous frequency-amplitude modulation. The empirical mode corresponding to the VM BP profile was used to construct the continuous phase diagram and to identify the minimum and maximum values from the residual BP (BP_R) and BFV (BFV_R) signals. The BP-BFV phase shift was calculated as the difference between the phase corresponding to the BP_R and BFV_R minimum (maximum) values. BP-BFV phase shifts were significantly different between groups. In the normotensive group, the BFV_R minimum and maximum preceded the BP_R minimum and maximum, respectively, leading to large positive values of BP-BFV shifts.

Conclusion

In the stroke and hypertensive groups, the resulting BP-BFV phase shift was significantly smaller compared to the normotensive group. A standard autoregulation index did not differentiate the groups. The MMPF method enables evaluation of autoregulatory dynamics based on instantaneous BP-BFV phase analysis. Regulation of BP-BFV dynamics is altered with hypertension and after stroke, rendering blood flow dependent on blood pressure.