Gait Speed and Grip Strength Are Associated With Dropping Out of the Liver Transplant Waiting List.

INTRODUCTION: Frailty measures can predict perioperative surgical risk in liver transplant patients. The 5-meter walk test (5MWT) and hand grip strength (HGS) are easy and reproducible frailty measures. We hypothesized that they could capture frailty in liver transplant listed patients and would be associated with dropping out of the waiting list. METHODS: We conducted a retrospective analysis of patients undergoing outpatient liver transplant listing at the University of Pittsburgh Medical Center from 2013 to 2016. We compared demographics, baseline laboratory markers, 5MWT, and HGS between patients who were dropped from the waiting list for medical reasons and those who remained or were successfully transplanted. Bivariate statistical analysis was performed using Fisher exact or χ2 tests. RESULTS: We reviewed 197 patients listed for liver transplant. Average age was 57.1 years (range 20-74), and patients were predominantly white (90.4%). Patients' most common etiology of liver disease was hepatitis C (32.5%), 14 (7.1%) had a previous liver transplant, and average Model for End-Stage Liver Disease score upon listing was 16.0. Of the cohort, 38 (19.3%) were ultimately dropped from the waitlist due to non-hepatocellular carcinoma-related reasons. Patients dropped from the waiting list had weaker HGS (46.14 lb vs 59.6 lb; P < .005) and slower 5MWT speed (5MWT: 0.92 m/s vs 1.03 m/s; P < .005). CONCLUSION: The 5MWT and HGS can easily measure frailty in patients being evaluated for liver transplant. These tests are associated with waiting list dropout, indicating that they can be valuable tools in the evaluation of these patients.

The Impact of Colleague Peer Review on the Radiotherapy Treatment Planning Process in the Radical Treatment of Lung Cancer.

AIMS: Modern radiotherapy uses techniques to reliably identify tumour and reduce target volume margins. However, this can potentially lead to an increased risk of geographic miss. One source of error is the accuracy of target volume delineation (TVD). Colleague peer review (CPR) of all curative-intent lung cancer plans has been mandatory in our institution since May 2013. At least two clinical oncologists review plans, checking treatment paradigm, TVD, prescription dose tumour and critical organ tolerances. We report the impact of CPR in our institution. MATERIALS AND METHODS: Radiotherapy treatment plans of all patients receiving radical radiotherapy were presented at weekly CPR meetings after their target volumes were reviewed and signed off by the treating consultant. All cases and any resultant change to TVD (including organs at risk) or treatment intent were recorded in our prospective CPR database. The impact of CPR over a 13 month period from May 2013 to June 2014 is reported. RESULTS: One hundred and twenty-two patients (63% non-small cell lung carcinoma, 17% small cell lung carcinoma and 20% 'clinical diagnosis') were analysed. On average, 3.2 cases were discussed per meeting (range 1-8). CPR resulted in a change in treatment paradigm in 3% (one patient proceeded to induction chemotherapy, two patients had high-dose palliative radiotherapy). Twenty-one (17%) had a change in TVD and one (1%) patient had a change in dose prescription. In total, 6% of patients had plan adjustment after review of dose volume histogram. CONCLUSION: The introduction of CPR in our centre has resulted in a change in a component of the treatment plan for 27% of patients receiving curative-intent lung radiotherapy. We recommend CPR as a mandatory quality assurance step in the planning process of all radical lung plans.

Production of a polyclonal anti-myostatin antibody and the effects of in ovo administration of the antibody on posthatch broiler growth and muscle mass.

In this study, we produced a polyclonal antibody against unprocessed chicken myostatin and examined the effect of in ovo administration of the antibody on posthatch chicken growth and muscle mass. A PCR-amplified unprocessed chicken myostatin cDNA was cloned into an Escherichia coli expression vector, and myostatin proteins were expressed. Recombinant myostatin purified by electro-elution of the SDS-PAGE fractionated myostatin band was used as an immunogen to produce rabbit polyclonal antimyostatin antibody (pAb-AVM46). In Western blot analysis, the pAb-AVM46 showed high affinity to the myostatin propeptide, but little affinity to the mature myostatin. Two experiments examined the effect of in ovo administration of the pAb-AVM46 on posthatch chicken growth and skeletal muscle mass. In experiment 1, broilers from eggs injected once with 35 microg of the antibody into the yolk on d 3 of incubation had significantly lower combined thigh and leg weight at 4 wk posthatch than the controls that received no injection, or the broilers from eggs received the same dose of antibody into the albumen. In experiment 2, 2 different doses of the antibody (9 or 70 microg) were injected into the yolk, and the effects on body and muscle weight were examined at 5 wk posthatch. Birds from eggs injected with 70 microg of the antibody had significantly lighter (11.6%) combined thigh and leg weight than the control birds. The percentage of the combined thigh and leg weight to BW of the 70-microg group was also significantly lower than that of the control group (20.95 vs. 23.08%). The results of this study indicate that unprocessed full-length myostatin as an immunogen produced antibody populations having affinity mostly to the propeptide with little to the mature form. The decreased muscle weight observed in broilers injected with the antibody in the yolk indicates that myostatin activity was probably elevated by the binding of the antibody to the propeptide, and provides evidence that myostatin propeptide inhibits the biological activity of myostatin in broilers.

Identification of a novel low-temperature-response element in the promoter of the barley (Hordeum vulgare L) gene blt101.1.

Two winter barley (Hordeum vulgare L. cv. Igri) genomic clones, lambda gblt101.1 and lambda gblt101.2, encoding the blt101 gene family, were isolated from a genomic library. Deletion analysis of the blt101.1 promoter, using transient beta-glucuronidase (GUS) reporter expression assays, indicated that it contains at least three regulatory regions. A 107-bp region between nucleotides -168 and -275 with respect to the translation initiation codon, confers high-level GUS reporter expression at low temperature and contains a sequence (designated CR1) that is highly conserved in equivalent positions within the promoters of both members of the blt101 gene family. A 10-bp motif contained within CR1 binds proteins present in nuclear extracts from both control and low-temperature-treated barley tissue. Loss-of-function experiments, using transient-expression analysis, confirmed that this motif acts as a previously unreported low-temperature-responsive element. Nuclease sensitivity analysis of intact chromatin indicated that the blt101.1 promoter becomes more susceptible to DNase and micrococcal nuclease at low temperature, consistent with chromatin reorganisation upon transcriptional induction. It is proposed that both the 10-bp motif and chromatin reorganisation are involved in the regulation of blt101.1 at low temperature. This is the first detailed analysis of a low-temperature-specific plant promoter and identifies a novel low-temperature-response element.

Aluminum toxicity alters the regulation of calbindin-D28k protein and mRNA expression in chick intestine.

Previous studies have shown that aluminum inhibits vitamin D-dependent calcium absorption. The mechanism involves reduced sensitivity to 1,25-dihydroxycholecalciferol and reduced expression of the calcium transport protein, calbindin-D28k. Reduced expression of calbindin protein may be due to decreased levels of calbindin mRNA. To test this hypothesis, we measured calbindin mRNA levels in chicks fed diets with and without added aluminum. Groups of chicks were fed one of four diets: control, control plus aluminum, low calcium, or low calcium plus aluminum. A fifth group was fed a vitamin D-free diet as a negative control. Calbindin protein was measured by immunoblotting. Serum calcium and inorganic phosphorus were determined. Intestinal mRNA was isolated and assayed by slot-blot hybridization to a fluorescein-conjugated oligonucleotide probe complementary to calbindin-D28k mRNA. Antifluorescein antibodies conjugated to alkaline phosphatase were used to detect hybrids and mRNA levels were quantified by densitometry. Specificity of the probe was verified by Northern analysis. Intestinal calbindin protein was greater in the control plus aluminum group than in controls, but no difference in calbindin mRNA was observed. These changes were associated with small decreases in serum phosphorus and calcium, suggesting a postranscriptional effect of aluminum. Chicks fed the low calcium diet had greater intestinal calbindin protein and mRNA levels relative to the control group in association with a 45% decrease in serum calcium. In contrast, no difference in calbindin protein, and significantly less mRNA were found in the low calcium plus aluminum group compared with controls, despite a decrease in serum calcium similar to that of chicks fed the low calcium diet without aluminum. These results show that in chicks fed a low calcium diet, aluminum intake decreases transcription and/or stability of intestinal calbindin mRNA, and that aluminum may inhibit the expression of vitamin D-dependent genes.

A review of two alternative ultrasound quality assurance programmes.

OBJECTIVE: This study reviewed the results of B-mode Quality Assurance (QA) performance tests on 17 real-time ultrasound scanners, performed over a period of 3 years, in order to assess their value. Following this review we revised and simplified our testing schedules to include two tests for noise and sensitivity. The value of the new schedules was assessed. METHODS: Initially, testing schedules were similar to those recommended by two professional bodies. Results were reviewed to determine whether the tests predicted or confirmed faults. We then introduced a simplified testing programme using alternative measurements, attempting to demonstrate or predict noise related faults that affect the image, but were not demonstrated by current tests. These new tests have been performed on 24 ultrasound machines for up to 18 months. RESULTS: A review of results has shown that measurements occasionally fall outside tolerance due to chance, and that faults that significantly affect the image, e.g. probe faults and noise, are reported by the users without predictive or concomitant changes in test results using our original schedules. Faults occur that do not immediately affect image quality and are not reported by the users. Inappropriate settings, e.g. monitors, are frequently reset at QA, particularly where there are potentially untrained users. The additional tests showed consistent changes in noise (four) or sensitivity (one) on five machines. CONCLUSION: Our earlier tests were inadequate in demonstrating deterioration in the clinical performance of ultrasound imaging equipment. Introduction of a revised testing schedule has resulted in changes in equipment performance being detected and rectified.

PIE-1 is a bifunctional protein that regulates maternal and zygotic gene expression in the embryonic germ line of Caenorhabditis elegans.

The CCCH zinc finger protein PIE-1 is an essential regulator of germ cell fate that segregates with the germ lineage during the first cleavages of the Caenorhabditis elegans embryo. We have shown previously that one function of PIE-1 is to inhibit mRNA transcription. Here we show that PIE-1 has a second function in germ cells; it is required for efficient expression of the maternally encoded Nanos homolog NOS-2. This second function is genetically separable from PIE-1's inhibitory effect on transcription. A mutation in PIE-1's second CCCH finger reduces NOS-2 expression without affecting transcriptional repression and causes primordial germ cells to stray away from the somatic gonad, occasionally exiting the embryo entirely. Our results indicate that PIE-1 promotes germ cell fate by two independent mechanisms as follows: (1) inhibition of transcription, which blocks zygotic programs that drive somatic development, and (2) activation of protein expression from nos-2 and possibly other maternal RNAs, which promotes primordial germ cell development.

Identification of essential active-site residues in the cyanogenic beta-glucosidase (linamarase) from cassava (Manihot esculenta Crantz) by site-directed mutagenesis.

The coding sequence of the mature cyanogenic beta-glucosidase (beta-glucoside glucohydrolase, EC 3.2.1.21; linamarase) was cloned into the vector pYX243 modified to contain the SUC2 yeast secretion signal sequence and expressed in Saccharomyces cerevisiae. The recombinant enzyme is active, glycosylated and showed similar stability to the plant protein. Michaelis constants for hydrolysis of the natural substrate, linamarin (K(m)=1.06 mM) and the synthetic p-nitrophenyl beta-D-glucopyranoside (PNP-Glc; K(m)=0.36 mM), as well as apparent pK(a) values of the free enzyme and the enzyme-substrate complexes (pK(E)(1)=4.4-4.8, pK(E)(2)=6.7-7.2, pK(ES)(1)=3.9-4.4, pK(ES)(2)=8.3) were very similar to those of the plant enzyme. Site-directed mutagenesis was carried out to study the function of active-site residues based on a homology model generated for the enzyme using the MODELLER program. Changing Glu-413 to Gly destroyed enzyme activity, consistent with it being the catalytic nucleophile. The Gln-339Glu mutation also abolished activity, confirming a function in positioning the catalytic diad. The Ala-201Val mutation shifted the pK(a) of the acid/base catalyst Glu-198 from 7.22 to 7.44, reflecting a change in its hydrophobic environment. A Phe-269Asn change increased K(m) for linamarin hydrolysis 16-fold (16.1 mM) and that for PNP-Glc only 2.5-fold (0.84 mM), demonstrating that Phe-269 contributes to the cyanogenic specificity of the cassava beta-glucosidase.

Asymmetric segregation of PIE-1 in C. elegans is mediated by two complementary mechanisms that act through separate PIE-1 protein domains.

The CCCH finger protein PIE-1 is a regulator of germ cell fate that segregates with the germ lineage in early embryos. At each asymmetric division, PIE-1 is inherited preferentially by the germline daughter and is excluded from the somatic daughter. We show that this asymmetry is regulated at the protein level by two complementary mechanisms. The first acts before cell division to enrich PIE-1 in the cytoplasm destined for the germline daughter. The second acts after cell division to eliminate any PIE-1 left in the somatic daughter. The latter mechanism depends on PIE-1's first CCCH finger (ZF1), which targets PIE-1 for degradation in somatic blastomeres. ZF1s in two other germline proteins, POS-1 and MEX-1, are also degraded in somatic blastomeres, suggesting that localized degradation also acts on these proteins to exclude them from somatic lineages.

Effect of dietary aluminum on tissue nonheme iron and ferritin levels in the chick.

Aluminum toxicity is well documented but the mechanism of action is poorly understood. In renal failure patients with aluminum overload, disturbances in iron metabolism leading to anemia are apparent. Few animal models, however, have been used to study the effects of dietary aluminum on iron metabolism. The purpose of this study was to determine if dietary aluminum exposure alters tissue iron and ferritin concentrations in the chick, as has been found in cultured human cells exposed to aluminum. Groups of day-old chicks were fed purified diets containing one of two levels of iron (control or high iron), and one of three levels of aluminum chloride in a 2 x 3 factorial design. Diets were consumed ad libitum for 1 week, then pair-feeding was initiated for 2 more weeks. A seventh group consumed a low iron diet ad libitum for comparative purposes. After the 3-week feeding period, samples of kidney, liver, and intestinal mucosa were analyzed for nonheme iron and ferritin concentrations by a colorimetric assay and SDS-PAGE, respectively. Results showed that dietary aluminum intake reduced iron stores in liver and intestine, but had no effect on nonheme iron levels in the kidney. Ferritin levels were reduced by aluminum intake in all tissues studied. The decreases in tissue ferritin levels were proportionately more than the decreases in tissue nonheme iron levels. This resulted in increased nonheme iron to ferritin ratios that amounted to as much as 140 and 525% in kidney and intestine, respectively. These findings are consistent with the interpretation that, in the growing chick, dietary aluminum can inhibit iron absorption, disrupt the regulation of tissue ferritin levels by iron, and potentially alter the compartmentalization and protective sequestration of iron within cells.

Transcriptional repression by the Caenorhabditis elegans germ-line protein PIE-1.

In the early Caenorhabditis elegans embryo, maternally expressed PIE-1 protein is required in germ-line blastomeres to inhibit somatic differentiation, maintain an absence of mRNA transcription, and block phosphorylation of the RNA polymerase II large subunit (Pol II) carboxy-terminal domain (CTD). We have determined that PIE-1 can function as a transcriptional repressor in cell culture assays. By fusing PIE-1 sequences to the yeast GAL4 DNA-binding domain, we have identified a PIE-1 repression domain that appears to inhibit the transcriptional machinery directly. A sequence element that is required for this repressor activity is similar to the Pol II CTD heptapeptide repeat, suggesting that the PIE-1 repression domain might target a protein complex that can bind the CTD. An alteration of this sequence element that blocks repression also impairs the ability of a transgene to rescue a pie-1 mutation, suggesting that this repressor activity may be important for PIE-1 function in vivo.

Identification of promoter elements in a low-temperature-responsive gene (blt4.9) from barley (Hordeum vulgare L.).

The blt4 barley gene family encodes non-specific lipid transfer proteins and has been shown, by in situ localisation, to be expressed in the epidermal cells of leaves. The transcriptionally controlled, low-temperature-responsive member of this gene family, blt4.9, is predominantly expressed in shoot meristems. The promoter region (1938 bp) of blt4.9 contains sequence motifs which have been implicated in responses to low temperature, abscisic acid and other environmental factors. Deletion analysis showed that a 42 bp sequence proximal to, but not including, the CAAT and TATA boxes, confers enhanced low-temperature response to a reporter gene in a barley shoot explant transient expression system. Other promoter regions were shown to contain negative and positive regulatory regions. Electrophoretic mobility shift analysis (EMSA) was used with nuclear proteins from either low-temperature- or control-temperature-treated plants to further investigate the blt4.9 promoter. Synthetic oligonucleotides were used to identify a hexanucleotide, CCGAAA, within the 42 bp, low-temperature-responsive promoter region, as the binding site of a low-mobility nuclear protein complex. This complex was present in nuclear extracts from both low-temperature-treated and control plants and was the only complex formed within this region. Mutation of the CCGAAA motif within the low-temperature-responsive 42 bp promoter sequence reduced low-temperature responsiveness to basal levels. A related upstream element, CCGAC, known to be a low-temperature-responsive element in other plants, did not bind to nuclear proteins in this study. It is proposed that the hexanucleotide CCGAAA, at -195 from the first ATG, is involved in the low-temperature response of blt4.9 in barley.

Transcriptionally repressed germ cells lack a subpopulation of phosphorylated RNA polymerase II in early embryos of Caenorhabditis elegans and Drosophila melanogaster.

Early embryonic germ cells in C. elegans and D. melanogaster fail to express many messenger RNAs expressed in somatic cells. In contrast, we find that ribosomal RNAs are expressed in both cell types. We show that this deficiency in mRNA production correlates with the absence of a specific phosphoepitope on the carboxy-terminal domain of RNA polymerase II. In both C. elegans and Drosophila embryos, this phosphoepitope appears in somatic nuclei coincident with the onset of embryonic transcription, but remains absent from germ cells until these cells associate with the gut primordium during gastrulation. In contrast, a second distinct RNA polymerase II phosphoepitope is present continuously in both somatic and germ cells. The germ-line-specific factor PIE-1 is required to block mRNA production in the germ lineage of early C. elegans embryos (Seydoux, G., Mello, C. C., Pettitt, J., Wood, W. B., Priess, J. R. and Fire, A. (1996) Nature 382, 713-716). We show here that PIE-1 is also required for the germ-line-specific pattern of RNA polymerase II phosphorylation. These observations link inhibition of mRNA production in embryonic germ cells to a specific modification in the phosphorylation pattern of RNA polymerase II and suggest that repression of RNA polymerase II activity may be part of an evolutionarily conserved mechanism that distinguishes germ line from soma during early embryogenesis. In addition, these studies also suggest that different phosphorylated isoforms of RNA polymerase II perform distinct functions.

mRNA stability and localisation of the low-temperature-responsive barley gene family blt14.

Transcription and translation inhibitors have been used to investigate the role of mRNA stability in the low-temperature-regulated expression of the post-transcriptionally controlled low temperature responsive barley gene family, blt14. Genomic clones (blt14.1, blt14.2) representing additional members of the blt14 gene family have been isolated and sequenced. Gene specific probes have been used to analyse the spatial expression of each individual member of the blt14 gene family. Findings indicate that all of the genes are responsive to low temperature, but the organ distribution is different for each gene. The results indicate that blt14.0 mRNA is stabilised by a low-temperature-dependent protein factor. Taken together, the results suggest that organ-specific post-transcriptional mechanisms are important in the low-temperature regulation of blt14 gene expression.

A low-temperature-responsive gene from barley encodes a protein with single-stranded nucleic acid-binding activity which is phosphorylated in vitro.

A low-temperature-responsive gene, blt 801, isolated from a winter barley (Hordeum vulgare L.) cDNA library prepared from leaf meristematic tissue, was sequenced. The deduced amino acid sequence predicts a glycine-rich RNA-binding protein (GR-RNP) which was homology to stress-responsive GR-RNPs from several other plant species. BLT 801 is a two-domain protein, the amino-terminal domain comprises a consensus RNA-binding domain similar to that found in many eukaryotic genes and the carboxy-terminal domain is extremely glycine-rich (68.5% glycine). Blt 801 mRNA also accumulates in response to the phytohormone abscisic acid. The protein encoded by blt 801 has been produced as a recombinant fusion protein using a bacterial expression vector. The fusion protein, a chimaera of glutathione S-transferase and BLT 801, has been used in studies to determine nucleic acid binding and other characteristics. Binding studies with single-stranded nucleic acids show that BLT 801 has affinity for homoribopolymers G, A and U but not C, it also binds to single-stranded DNA and selects RNA molecules containing open loop structures enriched in adenine but low in cytosine. Blt 801 has a consensus motif for phosphorylation by cAMP protein kinase (PKA) at the junction between the two domains which can be phosphorylated by PKA in vitro and which, by analogy to animal studies, may have significance for controlling enzyme function.

Lack of X inactivation associated with maternal X isodisomy: evidence for a counting mechanism prior to X inactivation during human embryogenesis.

We have previously reported functional disomy for X-linked genes in females with tiny ring X chromosomes and a phenotype significantly more abnormal than Turner syndrome. In such cases the disomy results from failure of these X chromosomes to inactivate because they lack DNA sequences essential for cis X inactivation. Here we describe a novel molecular mechanism for functional X disomy that is associated with maternal isodisomy. In this case, the severe mental retardation and multiple congenital abnormalities in a female with a mosaic 45,X/ 46,X,del(X)(q21.3-qter)/ 46X,r(X) karyotype are associated with overexpression of the genes within Xpter to Xq21.31 in many of her cells. Her normal X, ring X, and deleted linear X chromosomes originate from the same maternal X chromosome, and all are transcriptionally active. None expresses X inactive specific transcript (XIST), although the locus and region of the putative X inactivation center (XIC) are present on both normal and linear deleted X chromosomes. To our knowledge, this is the first report of a functional maternal X isodisomy, and the largest X chromosome to escape inactivation. In addition, these results (1) show that cis inactivation does not invariably occur in human females with two X chromosomes, even when the XIC region is present on both of them; (2) provide evidence for a critical time prior to the visible onset of X inactivation in the embryo when decisions about X inactivation are made; and (3) support the hypothesis that the X chromosome counting mechanism involves chromosomal imprinting, occurs prior to the onset of random inactivation, and is required for subsequent inactivation of the chromosome.

Characterization of a novel protein in chick intestine that exhibits calcium-binding activity and regulation by dietary calcium, aluminum and vitamin D.

The molecular mechanisms stimulated by vitamin D and low calcium diets that promote intestinal calcium absorption are not fully understood. In the present experiments, groups of chicks were subjected to the following treatments known to alter the efficiency of Ca absorption: vitamin D deficiency, repletion of vitamin D-deficient chicks with 1,25-dihydroxycholecalciferol [1,25(OH)2D3], low Ca intakes, and aluminum toxicity. Duodenal mucosal scrapings were obtained and screened for changes in protein composition using SDS-PAGE and size exclusion chromatography. The relative amount of a previously unreported 400-kDa oligomer containing 22-kDa monomeric subunits was found to vary directly with theoretical changes in the efficiency of Ca absorption, i.e., amounts increased with low Ca intakes and repletion with 1,25(OH)2D3, but were decreased by dietary aluminum and vitamin D deficiency. The oligomer was shown to have Ca-binding activity using a 45Ca overlay technique. These properties suggest 1) that the protein is regulated, at least indirectly, by 1,25(OH)2D3; 2) that the protein may play a role in promoting Ca absorption, possibly by binding Ca; and 3) that dietary aluminum interferes with the regulation of this protein, possibly by interfering with the actions of vitamin D in the intestine.

Electrophysiologic manifestations of impaired temporal lobe auditory processing in verbal auditory agnosia.

The present study examined the extent to which verbal auditory agnosia (VAA) is primarily a phonemic decoding disorder, as contrasted to a more global defect in acoustic processing. Subjects were six young adults who presented with VAA in childhood and who, at the time of testing, showed varying degrees of residual auditory discrimination impairment. They were compared to a group of young adults with normal language development matched for age and gender. Cortical event-related potentials (ERPs) were recorded to tones and to consonant-vowel stimuli presented in an "oddball" discrimination paradigm. In addition to cortical ERPs, auditory brainstem responses (ABRs) and middle latency responses (MLRs) were recorded. Cognitive and language assessments were obtained for the VAA subjects. ABRs and MLRs were normal. In comparison with the control group, the cortical ERPs of the VAA subjects showed a delay in the N1 component recorded over lateral temporal cortex both to tones and to speech sounds, despite an N1 of normal latency overlying the frontocentral region of the scalp. These electrophysiologic findings indicate a slowing of processing of both speech and nonspeech auditory stimuli and suggest that the locus of this abnormality is within the secondary auditory cortex in the lateral surface of the temporal lobes.

Studies of X inactivation and isodisomy in twins provide further evidence that the X chromosome is not involved in Rett syndrome.

Rett syndrome (RS), a progressive encephalopathy with onset in infancy, has been attributed to an X-linked mutation, mainly on the basis of its occurrence almost exclusively in females and its concordance in female MZ twins. The underlying mechanisms proposed are an X-linked dominant mutation with male lethality, uniparental disomy of the X chromosome, and/or some disturbance in the process of X inactivation leading to unequal distributions of cells expressing maternal or paternal alleles (referred to as a "nonrandom" or "skewed" pattern of X inactivation). To determine if the X chromosome is in fact involved in RS, we studied a group of affected females including three pairs of MZ twins, two concordant for RS and one uniquely discordant for RS. Analysis of X-inactivation patterns confirms the frequent nonrandom X inactivation previously observed in MZ twins but indicates that this is independent of RS. Analysis of 29 RS females reveals not one instance of uniparental X disomy, extending the observations previously reported. Therefore, our findings contribute no support for the hypothesis that RS is an X-linked disorder. Furthermore, the concordant phenotype in most MZ female twins with RS, which has not been observed in female twins with known X-linked mutations, argues against an X mutation.

Reversal of advanced liver fibrosis in rabbits with Schistosomiasis japonica.

Advanced liver fibrosis is generally considered to be irreversible. We studied the reversibility of marked liver fibrosis in rabbits infected with Schistosoma japonicum. We determined liver collagen content, collagen biosynthesis, and collagenase activity using serial biopsy specimens obtained 20, 40, and 60 weeks after infection. Reversibility of this process was investigated in rabbits cured of infection at 21 weeks; control rabbits not cured of infection were also studied. At 20 weeks, liver collagen content was 16-fold greater than normal, with accumulation of collagen types I, III, and V. Synthesis of collagen within fibrotic liver slices was 10-fold greater than normal. Liver collagenolytic activity for a type I substrate was 19-fold greater than normal. After parasitologic cure, a striking morphologic reversal of fibrosis occurred during the subsequent 40 weeks, with the return of liver collagen content to three-fold greater than normal and a 75% decrease in synthetic rates compared with those at 20 weeks (P < 0.01). Collagenolytic activity remained elevated to the same degree noted at 20 weeks. A similar but lesser resolution of fibrosis also occurred in untreated control rabbits, coincident with a spontaneous decrease in new egg deposition known to occur in this model system. We conclude that advanced liver fibrosis in S. japonicum-infected rabbits is slowly reversible after cure or senescence of the infection. A possible mechanism for this reversal is persistently increased collagenolysis as collagen synthesis diminishes.