Expression of Reg/PAP family members during motor nerve regeneration in rat

In this study, we examined the expression of mRNAs for Regenerating gene (Reg)/pancreatitis-associated protein (PAP) family members following hypoglossal nerve injury in rats. In addition to four rat family members (RegI, Reg-2/PAP I, PAP II, and PAP III) that had been identified, we newly cloned and sequenced a type-IV Reg gene in rats. Among these five family members, the expression of Reg-2/PAP I mRNA was predominantly enhanced in injured motor neurons after axotomy. Furthermore, a marked induction of PAP III mRNA was observed in the distal part of the injured nerve. A polyclonal antibody was raised against PAP III, and a Western blotting analysis using this antibody confirmed an increased level of PAP III protein in the injured nerve. These results suggest that Reg family members would be new mediators among injured neurons and glial cells, and may play pivotal roles during nerve regeneration.     

Rat kinesin light chain 3 associates with spermatid mitochondria

We recently discovered that in rat spermatids, kinesin light chain KLC3 can associate with outer dense fibers, major sperm tail components, and accumulates in the sperm midpiece. Here, we show that mitochondria isolated from rat-elongating spermatids have bound KLC3. Immunoelectron microscopy indicates that the association of KLC3 with mitochondria coincides with the stage in spermatogenesis when mitochondria move from the plasma membrane to the developing midpiece. KLC3 is able to bind in vitro to mitochondria from spermatids as well as somatic cells employing a conserved kinesin light chain motif, the tetratrico-peptide repeats. Expression of KLC3 in fibroblasts results in formation of large KLC3 clusters close to the nucleus, which also contain mitochondria: no other organelles were present in these clusters. Mitochondria are not present in KLC3 clusters after deletion of KLC3's tetratrico-peptide repeats. Our results indicate that the rat spermatid kinesin light chain KLC3 can associate with mitochondria.     

Estimating the cumulative risk of a false-positive test in a repeated screening program

The goal of screening tests for a chronic disease such as cancer is early detection and treatment with a consequent reduction in mortality from the disease. Screening tests, however, might produce false positive and false-negative results. With an increasing number of screening tests, it is clear that the risk of a false-positive screen, a finding with potentially significant emotional, financial, and health costs, also increases. Elmore et al. (1998, New England Journal of Medicine 338, 1089-1096), Christiansen et al. (2000, Journal of the National Cancer Institute 92, 1657-1666), and Gelfand and Wang (2000, Statistics in Medicine 19, 1865-1879) investigated this problem under the somewhat unrealistic assumption that the choice of making the decision to drop out at the kth screen does not depend upon the results of the earlier k - 1 screens. In this article we obtain sufficient and necessary conditions for their assumption to hold and use one of them to provide a method for testing the validity of the assumption. A new model which does not depend on their assumption is introduced. The maximum likelihood estimator of the cumulative risk of receiving a false-positive screen under the new model is derived and its asymptotic normality is proved. The extension of the new model by incorporating covariate information is also considered. We apply our testing method and the new model to data from the breast cancer screening trial of the Health Insurance Plan of Greater New York.     

Involvement of caspase-9 in execution of the maternal program of apoptosis in Xenopus late blastulae overexpressed with S-adenosylmethionine decarboxylase

We previously demonstrated that overexpression of S-adenosylmethionine decarboxylase (SAMDC) in Xenopus early embryos induces execution of maternal program of apoptosis shortly after midblastula transition, which likely serves as a fail-safe mechanism of early development to eliminate physiologically damaged cells before they entering the gastrula stage. To determine how caspases are involved in this process, we microinjected peptide inhibitors and "dominant-negative forms" of caspase-9 and -1 into Xenopus fertilized eggs, and found that inhibitors of caspase-9, but not caspase-1, completely suppress SAMDC-induced apoptosis. The lysate of SAMDC-overexpressing late blastulae contained activity to cleave in vitro-synthesized [(35)S]procaspase-9, but not [(35)S]procaspase-1, and mRNA for caspase-9, but not caspase-1, occurred abundantly in the unfertilized egg as maternal mRNA. We also found that overexpression of caspase-9 and -1 equally executes the apoptosis, but the apoptosis executed by these mRNAs was only partially rescued by Bcl-2 and rescued embryos did not develop beyond neurula stage. These results indicate that activation of caspase-9 is a key step for execution of the maternally preset program of apoptosis in Xenopus early embryos.     

G-IMEx: A comprehensive software tool for detection of microsatellites from genome sequences

Microsatellites are ubiquitous short tandem repeats found in all known genomes and are known to play a very important role in various studies and fields including DNA fingerprinting, paternity studies, evolutionary studies, virulence and adaptation of certain bacteria and viruses etc. Due to the sequencing of several genomes and the availability of enormous amounts of sequence data during the past few years, computational studies of microsatellites are of interest for many researchers. In this context, we developed a software tool called Imperfect Microsatellite Extractor (IMEx), to extract perfect, imperfect and compound microsatellites from genome sequences along with their complete statistics. Recently we developed a user-friendly graphical-interface using JAVA for IMEx to be used as a stand-alone software named G-IMEx. G-IMEx takes a nucleotide sequence as an input and the results are produced in both html and text formats. The Linux version of G-IMEx can be downloaded for free from http://www.cdfd.org.in/imex.     

The Legs at odd angles (Loa) Mutation in Cytoplasmic Dynein Ameliorates Mitochondrial Function in SOD1G93A Mouse Model for Motor Neuron Disease

Amyotrophic lateral sclerosis (ALS) is a debilitating and fatal late-onset neurodegenerative disease. Familial cases of ALS (FALS) constitute ∼10% of all ALS cases, and mutant superoxide dismutase 1 (SOD1) is found in 15–20% of FALS. SOD1 mutations confer a toxic gain of unknown function to the protein that specifically targets the motor neurons in the cortex and the spinal cord. We have previously shown that the autosomal dominant Legs at odd angles (Loa) mutation in cytoplasmic dynein heavy chain (Dync1h1) delays disease onset and extends the life span of transgenic mice harboring human mutant SOD1^G93A. In this study we provide evidence that despite the lack of direct interactions between mutant SOD1 and either mutant or wild-type cytoplasmic dynein, the Loa mutation confers significant reductions in the amount of mutant SOD1 protein in the mitochondrial matrix. Moreover, we show that the Loa mutation ameliorates defects in mitochondrial respiration and membrane potential observed in SOD1^G93A motor neuron mitochondria. These data suggest that the Loa mutation reduces the vulnerability of mitochondria to the toxic effects of mutant SOD1, leading to improved mitochondrial function in SOD1^G93A motor neurons.     

Paired-Pulse Facilitation of Transmitter Release at Different Levels of Extracellular Calcium Concentration

High-frequency synaptic activity can cause facilitation of transmitter release due to accumulation of “residual Ca²⁺” at the nerve terminal. However, the mechanism of this phenomenon is still under debate. Here we show that, using extracellular recording from frog cutaneous pectoris muscle, paired-pulse facilitation (PPF) at the frog neuro-muscular junction decays in two or three-exponential manner depending upon the extracellular Ca²⁺ concentration ([Ca²⁺]_e). First, second and “early” PPF components are analyzed and described in this study. Considering the dependence of PPF on [Ca²⁺]_e, existence of several specific high-affinity intra-terminal Ca²⁺-binding sites that underlie the facilitation of transmitter release at the frog neuro-muscular junction is proposed.     

The role of ventral premotor cortex in action execution and action understanding

The human ventral premotor cortex overlaps, at least in part, with Broca's region in the dominant cerebral hemisphere, that is known to mediate the production of language and contributes to language comprehension. This region is constituted of Brodmann's areas 44 and 45 in the inferior frontal gyrus. We summarize the evidence that the motor related part of Broca's region is localized in the opercular portion of the inferior frontal cortex, mainly in area 44 of Brodmann. According to our own data, there seems to be a homology between Brodmann area 44 in humans and the monkey area F5. The non-language related motor functions of Broca's region comprise complex hand movements, associative sensorimotor learning and sensorimotor integration. Brodmann's area 44 is also a part of a specialized parieto-premotor network and interacts significantly with the neighbouring premotor areas. In the ventral premotor area F5 of monkeys, the so called mirror neurons have been found which discharge both when the animal performs a goal-directed hand action and when it observes another individual performing the same or a similar action. More recently, in the same area mirror neurons responding not only to the observation of mouth actions, but also to sounds characteristic to actions have been found. In humans, through an fMRI study, it has been shown that the observation of actions performed with the hand, the mouth and the foot leads to the activation of different sectors of Broca's area and premotor cortex, according to the effector involved in the observed action, following a somatotopic pattern which resembles the classical motor cortex homunculus. On the other hand the evidence is growing that human ventral premotor cortex, especially Brodmann's area 44, is involved in polymodal action processing. These results strongly support the existence of an execution-observation matching system (mirror neuron system). It has been proposed that this system is involved in polymodal action recognition and might represent a precursor of language processing. Experimental evidence in favour of this hypothesis both in the monkey and humans is shortly reviewed.     

Differential distribution of antibodies to different viruses in young animals in the free-ranging rhesus macaques of Cayo Santiago

BACKGROUND: The breeding colony of free-ranging rhesus macaques was established in 1938 in Cayo Santiago (CS) with animals collected in northern India. The seroprevalence to cercopithecine herpesvirus type 1 (B virus) and simian retroviruses has been studied previously. RESULTS: This is the first report on the seropositivity to different viruses using samples collected shortly after removing animals (n = 245) from CS. All samples were negative for measles, simian immunodeficiency virus and simian type D retroviruses. The overall prevalence of antibodies was around 50% for simian T-lymphotropic virus I (STLV-I). For B virus, the prevalence was 38%. CONCLUSIONS: Data obtained showed marked differences in the antibody distribution to B virus and STLV-I within the free-ranging colony of rhesus macaques. Implication of these data for the Specific Pathogen Free program at the Caribbean Primate Research Center are also discussed.