Omentum and bone marrow: how adipocyte-rich organs create tumour microenvironments conducive for metastatic progression.

A number of clinical studies have linked adiposity with increased cancer incidence, progression and metastasis, and adipose tissue is now being credited with both systemic and local effects on tumour development and survival. Adipocytes, a major component of benign adipose tissue, represent a significant source of lipids, cytokines and adipokines, and their presence in the tumour microenvironment substantially affects cellular trafficking, signalling and metabolism. Cancers that have a high predisposition to metastasize to the adipocyte-rich host organs are likely to be particularly affected by the presence of adipocytes. Although our understanding of how adipocytes influence tumour progression has grown significantly over the last several years, the mechanisms by which adipocytes regulate the metastatic niche are not well-understood. In this review, we focus on the omentum, a visceral white adipose tissue depot, and the bone, a depot for marrow adipose tissue, as two distinct adipocyte-rich organs that share common characteristic: they are both sites of significant metastatic growth. We highlight major differences in origin and function of each of these adipose depots and reveal potential common characteristics that make them environments that are attractive and conducive to secondary tumour growth. Special attention is given to how omental and marrow adipocytes modulate the tumour microenvironment by promoting angiogenesis, affecting immune cells and altering metabolism to support growth and survival of metastatic cancer cells.

ProLuCID: An improved SEQUEST-like algorithm with enhanced sensitivity and specificity.

ProLuCID, a new algorithm for peptide identification using tandem mass spectrometry and protein sequence databases has been developed. This algorithm uses a three tier scoring scheme. First, a binomial probability is used as a preliminary scoring scheme to select candidate peptides. The binomial probability scores generated by ProLuCID minimize molecular weight bias and are independent of database size. A modified cross-correlation score is calculated for each candidate peptide identified by the binomial probability. This cross-correlation scoring function models the isotopic distributions of fragment ions of candidate peptides which ultimately results in higher sensitivity and specificity than that obtained with the SEQUEST XCorr. Finally, ProLuCID uses the distribution of XCorr values for all of the selected candidate peptides to compute a Z score for the peptide hit with the highest XCorr. The ProLuCID Z score combines the discriminative power of XCorr and DeltaCN, the standard parameters for assessing the quality of the peptide identification using SEQUEST, and displays significant improvement in specificity over ProLuCID XCorr alone. ProLuCID is also able to take advantage of high resolution MS/MS spectra leading to further improvements in specificity when compared to low resolution tandem MS data. A comparison of filtered data searched with SEQUEST and ProLuCID using the same false discovery rate as estimated by a target-decoy database strategy, shows that ProLuCID was able to identify as many as 25% more proteins than SEQUEST. ProLuCID is implemented in Java and can be easily installed on a single computer or a computer cluster. This article is part of a Special Issue entitled: Computational Proteomics.

[Functional asplenia after severe varicella zoster-infection diagnosis by colour Doppler ultrasound]. / Autosplenektomie nach Varizellensepsis bei einem Patienten mit M. Crohn unter immunsuppressiver Therapie.

Infiltrative, inflammatory or thromboembolic processes in the parenchyma of the spleen can cause a functional loss of the organ. This phenomenon is called functional asplenia and occurs as a complication especially in sickle cell disease, lupus erythematosus and after bone marrow transplantation. We present the case of a patient with Crohn's disease under immunosuppressive therapy who developed a spontaneous covered spleen rupture in the course of a septic shock with DIG due to a Varizella zoster infection. Later on, sonography showed a diminution of the spleen size. No flow signals could be derived by colour doppler measurements from the spleen. Because of the colour doppler findings we suspected a functional asplenia which was then verified by spleen scintigraphy and Howell-Jolly-Bodies in the blood count. Remarkably, the Crohn's disease remains in complete remission since the development of the functional asplenia (for 4 years now). The underlying pathomechanism remains unclear.

Fluorescent oligonucleotide probes for clinical and environmental detection of Acanthamoeba and the T4 18S rRNA gene sequence type.

The first genus- and subgenus-specific fluorescent oligonucleotide probes for in situ staining of Acanthamoeba are described. Sequences of these phylogeny-based probes complement the 18S rRNA and the gene encoding it (18S rDNA). The genus-specific probe (GSP) is a fluorescein-labeled 22-mer specific for Acanthamoeba as shown here by its hybridization to growing trophozoites of all 12 known Acanthamoeba 18S rDNA sequence types and by its failure to hybridize with amoebae of two other genera (Hartmannella vermiformis and Balamuthia mandrillaris), two human cell lines, and two bacteria (Pseudomonas aeruginosa and Escherichia coli). The sequence type T4-specific probe (ST4P) is a rhodamine-labeled 30-mer specific for Acanthamoeba 18S rDNA sequence type T4, as shown here in hybridization tests with trophozoites of all 12 sequence types. T4 is the subgenus group associated most closely with Acanthamoeba keratitis (AK). GSP also was tested with corneal scrapings from 17 patients with a high index of clinical suspicion of AK plus 5 patient controls. GSP stained both trophozoites and cysts, although nonspecific cyst wall autofluorescence also was observed. Results could be obtained with GSP in 1 to 2 days, and based on results from cell culture tests, the probe correctly detected the presence or absence of Acanthamoeba in 21 of 24 specimens from the 22 patients. The use of GSP with cultured trophozoites and cysts from corneal scrapings has illustrated the suitability of using fluorescent oligonucleotide probes for identification of the genus Acanthamoeba in both environmental and clinical samples. In addition, the use of ST4P with cultured amoebae has indicated the potential of oligonucleotide probes for use in subgenus classification.

Acanthamoeba strains isolated from organs of freshwater fishes.

Contrary to data on Acanthamoeba infections in humans, little is known about infections in fishes. The present study combines the description of strains isolated from fishes with presentation of an improved method for subgeneric classification. Acanthamoeba spp. were isolated aseptically from tissues of 14 (1.7%) of 833 asymptomatic fishes collected in rivers and streams in the Czech Republic. Acanthamoebae successfully cloned from 10 of the 14 isolated strains were examined here. Morphology of these isolates was evaluated using light optics plus scanning and transmission electron microscopy. Cyst morphology, which varied extensively within and among clones, was most like morphological group II, but species-level classification was considered impossible. A distance analysis based on 442 bases in an 18S rDNA polymerase chain reaction fragment of about 460 bp placed the isolates in a clade composed of sequence types T3, T4, and T11, the 3 subdivisions of morphological group II. Fluorescent in situ hybridization (FISH) using oligonucleotide probes indicated that all isolates belong to a single subdivision of group II, the T4 sequence type. It has been concluded that the fish isolates are most closely related to strains commonly isolated from human infections, especially Acanthamoeba keratitis. The shorter diagnostic fragment sequences have proved nearly as useful as complete 18S rDNA sequences for identification of Acanthamoeba isolates.

A human papillomavirus related to human papillomavirus MM7/LVX82 produces distinct histological abnormalities in human foreskin implants grown as athymic mouse xenografts.

Studies of human papillomaviruses (HPVs) are hampered by the lack of a conventional culture system because HPV completes its life cycle only in fully differentiated human tissue. To overcome this obstacle, the athymic mouse xenograft system has been used to study the pathogenesis of HPV 11 and to develop neutralizing assays for vaccine development. Recently, HPV 40 has been produced in this system, and HPV 16 has been produced using mice with severe combined immune deficiency. To identify and characterize additional genital HPV types for similar studies, condylomata acuminata lesions containing a high copy number of HPV and detectable L1 major capsid protein were used to prepare infectious virus stocks. Human foreskin fragments were infected with the virus preparations and implanted under the renal capsules of athymic mice. After 5 months of growth, implant tissue was removed and processed for studies to detect HPV infection. Evidence of HPV infection was noted in some of the implants, but in contrast to HPV 11-infected epithelium, the implants derived from the new virus preparations contained a lesser degree of acanthosis, less developed koilocytosis, and a reduced number of preserved nuclei in the hyperkeratotic material within the cyst lining. The L1 consensus region was amplified by polymerase chain reaction from implant DNA and sequenced. Alignment of the amplified sequences with those in the HPV sequence database showed that the 452-bp amplimer was closely related but not identical to HPV LVX82 and HPV MM7 (also called Pap 291). The entire 7.9-kb genome was amplified by polymerase chain reaction and cloned. The presence of virions of the new isolate (named HPV IU) in the implants was verified by immunohistochemical detection of L1 major capsid protein and by demonstration of virion particles by electron microscopy. A second extract was made from one of the new implants and used to successfully propagate HPV IU. These experiments demonstrate that experimental infection of human epithelium with the new isolate, HPV IU, is associated with histological abnormalities that differ in potentially important ways from the changes observed in experimental HPV 11 infection.

Group-I introns with unusual sequences occur at three sites in nuclear 18S rRNA genes of Acanthamoeba lenticulata.

Seven of eleven isolates of Acanthamoeba lenticulata were found to have group-I introns located at one of three positions within the 18S rRNA gene. The introns are 636-721-bp long and are absent from mature rRNA. They lack open reading frames that could encode any known endonucleases. Sequences of introns from the same site in different isolates are 86.0-98.9% identical, while from different sites they are 24.2-29.8% identical. The most closely related introns from other organisms are in the 18S rRNA genes of several green algae where the 17.0-23.6% identity is mostly limited to a highly conserved core of base pairs including P, Q, R and S. Because the A. lenticulata introns only occur in one Acanthamoeba lineage, they were probably acquired after the divergence of this species.

A low concentration of ethanol reduces the chemiluminescence of human granulocytes and monocytes but not the tumor necrosis factor alpha production by monocytes after endotoxin stimulation.

The ability of polymorphonuclear neutrophils (PMNs) and monocytes (Mphi) to produce reactive oxygen species (ROS) has been related closely to their potential in the killing of microorganisms. Ethanol has been shown to impair the generation of ROS in these phagocytes after stimulation with some immunogens and to increase the susceptibility of alcohol abusers to infectious diseases. As endotoxemia is common in alcohol abusers, we investigated the effect of ethanol (21.7 mmol/liter) on the luminol-amplified chemiluminescence of PMNs and Mphi after endotoxin stimulation and the release of tumor necrosis factor alpha (TNF-alpha) from Mphi. Further, the efficiency of ethanol to inactivate chemically generated ROS was tested. Significant stimulation of ROS release occurred at endotoxin concentrations of 1 ng/ml or higher in both PMNs and Mphi. Ethanol significantly suppressed the formation of ROS in both cell types, the decrease being more pronounced in Mphi (-73. 8%) than in PMNs (-45.7%). The correlations between endotoxin concentration and the amount of released ROS showed a dose-dependent, sigmoidal course. Concentrations of endotoxin necessary for half-maximum stimulation were nearly identical (6 to 8 ng/ml) in both PMNs and Mphi, independent of the presence of ethanol. In contrast to ROS formation, ethanol had no effect on the amount of TNF-alpha produced by endotoxin-stimulated Mphi. Ethanol was shown to be unable to decrease the levels of chemically generated ROS under physiological conditions. Therefore, ethanol cannot be assumed to be an "antioxidative" compound but rather seems to modify processes of endotoxin recognition, intracellular signal transduction, or metabolism.

The evolutionary history of the genus Acanthamoeba and the identification of eight new 18S rRNA gene sequence types.

The 18S rRNA gene (Rns) phylogeny of Acanthamoeba is being investigated as a basis for improvements in the nomenclature and taxonomy of the genus. We previously analyzed Rns sequences from 18 isolates from morphological groups 2 and 3 and found that they fell into four distinct evolutionary lineages we called sequence types T1-T4. Here, we analyzed sequences from 53 isolates representing 16 species and including 35 new strains. Eight additional lineages (sequence types T5-T12) were identified. Four of the 12 sequence types included strains from more than one nominal species. Thus, sequence types could be equated with species in some cases or with complexes of closely related species in others. The largest complex, sequence type T4, which contained six closely related nominal species, included 24 of 25 keratitis isolates. Rns sequence variation was insufficient for full phylogenetic resolution of branching orders within this complex, but the mixing of species observed at terminal nodes confirmed that traditional classification of isolates has been inconsistent. One solution to this problem would be to equate sequence types and single species. Alternatively, additional molecular information will be required to reliably differentiate species within the complexes. Three sequence types of morphological group 1 species represented the earliest divergence in the history of the genus and, based on their genetic distinctiveness, are candidates for reclassification as one or more novel genera.

Risk factors for acute and subacute stent thrombosis after high-pressure stent implantation: a study by intracoronary ultrasound.

BACKGROUND: In the era of coronary stenting with high-pressure expansion, stent thrombosis remains a major life-threatening risk. Because of its superior imaging mode, intracoronary ultrasound could provide insight into potential risk factors for stent thrombosis. PATIENTS AND METHODS: In 215 patients with stent implantations controlled by intracoronary ultrasound, four occurrences of subacute stent thrombosis, two complete acute thrombotic occlusions, and four occurrences of incomplete acute thrombosis were observed. All stents were expanded with inflation pressures of at least 14 atm ( 17+/-3 atm). The clinical data and the qualitative and quantitative ultrasound information were compared between stents with thrombosis and stents without thrombosis. The luminal area and the plaque border of the reference segments, and of the smallest and largest site of the stented segment after the initial and final expansion, were measured. RESULTS: Stents in the left anterior descending artery were more often involved in stent thrombosis than other vessels, but the vessel dimensions in this target vessel were smaller than in the right coronary artery. The plaque burden was considerably larger after stent implantation with subsequent thrombosis compared with no thrombosis (74.1+/-8.8% vs 63.6+/-8.0%; p < 0.001), and the stent area was smaller (4.80%+/-1.33 mm2 vs 6.86+/-2.08 mm2; p < 0.01 ). In stents with thrombosis the plaque burden of the stent site with the smallest and largest lumen differed by 15.2%, whereas the difference in plaque burden in stents without thrombosis was 2.7%. Intracoronary ultrasound showed that the best risk predictor of thrombosis was the residual plaque burden of the stented segment (odds ratio 15.7 [confidence interval 2.4 to 104.7]), and a small stent area after implantation (odds ratio 6.8 [confidence interval 1.9 to 24.3]). CONCLUSION: In a multivariate risk analysis plaque burden was the strongest independent risk factor for stent thrombosis. The amount of residual plaque mass around the stent might be a potential trigger for thrombus formation.

Additional luminal area gain by intravascular ultrasound guidance after coronary stent implantation with high inflation pressure.

AIMS: Studies by intravascular ultrasound demonstrated inadequate expansion in a large number of stents, which lead to the increase of inflation pressure for stenting. The present study examined whether routine use of high-pressure inflation would be sufficient for an optimum stent expansion without sonographic guidance. METHODS AND RESULTS: Two types of single coronary stents (Palmaz-Schatz in 54, and Wiktor in 25) were implanted with inflation pressures of 16-20 atm in 79 nonocclusive coronary lesions. IVUS before stenting was used in 78% to select the adequate stent size. Intravascular ultrasound after stenting was used to asses the minimum stent are and diameter, the reference areas, and the strut apposition to the vessel wall. The difference between the area of the expanding balloon and the stent area was calculated as the luminal deficit of the stent. Completeness of stent expansion required full strut apposition and lesion coverage, and a minimum stent area that was larger than the distal reference, and larger than 60% of the proximal reference. Intravascular ultrasound before stenting lead to an increase of the stent size in 47%. After high-pressure expansion, even with the optimized balloon size, 8% of stents had struts protruding into the lumen. The stent area (6.87 +/- 1.93 mm2) was significantly smaller than both the proximal (9.59 +/- 2.91 mm2; p < 0.001) and distal reference area (8.23 +/- 3.03 mm2; p < 0.001). The criteria for complete expansion were met in 48%. The expansion with a larger high-pressure balloon in 28 stents lead to an increase of the stent area by 19% (8.19 +/- 2.24; p < 0.001), and full stent apposition in all cases. The criteria of stent expansion were met in 82%. A wide range of the luminal deficit upto 48% was observed, which was not related to sonographic lesion characteristics, except in lesions with complete circumferential calcifications. The different stent designs were characterized by a slightly lower luminal deficit in slotted-tube stents (23 +/- 13% vs. 28 +/- 12%; p = 0.11) and a better index of stent symmetry as compared with the coil stent (0.87 +/- 0.08 vs. 0.82 +/- 0.09; p < 0.05). CONCLUSION: Routine use of high-pressure stent expansion did not lead to a sufficient stent expansion, even when the initial stent size had been guided by intravascular ultrasound. Further stent dilatation with larger balloons under ultrasound guidance would be required to optimize the luminal area gain.

Causes of failed angioplasty for acute myocardial infarction assessed by intravascular ultrasound.

Coronary angioplasty is gaining increased importance as a primary treatment of acute myocardial infarction, but the complication rate of the procedure is higher than in stable coronary artery disease. In a consecutive series of 110 coronary angioplasties in patients with acute myocardial infarction, the cause of initially failed procedures was studied by intravascular ultrasound. The balloon angioplasty was immediately successful in 66%. In those cases with crossing of the lesion but a dissatisfying dilatation result (persistent occlusion, reocclusion, or dissection) an intravascular ultrasound probe could be advanced to 32 of 34 lesions. The information provided by ultrasound guided the subsequent bailout therapy. Persistent occlusions were caused by extensive thrombosis in 4.5% of all cases, subsequently treated by local thrombolysis for 12 to 16 hours, and in 1.8% by a ruptured plaque, which was treated by stenting. In cases with Thrombolysis in Myocardial Infarction (TIMI) flow II, angiography suggested a thrombus in 9.1%, but intravascular ultrasound could detect dissections instead of a thrombus in half the cases. In cases of dissection, stenting was performed. Dissections were observed by angiography in 15.5%, and all cases were confirmed by ultrasound. In vessels >2.5 mm the dissection was treated by stenting. Overall, in 20 of 21 lesions stents were successfully implanted. No stent thrombosis was observed. With the assistance of intravascular ultrasound during bailout therapy, the success of coronary angioplasty to achieve TIMI flow III without residual stenosis in an unselected consecutive patient cohort with acute myocardial infarction was 96%. In direct angioplasty for acute myocardial infarction the procedure is frequently complicated by events such as plaque rupture and extensive vascular thrombosis, which are uncommon in coronary angioplasty for stable angina. Intravascular ultrasound provided insight into the underlying morphologic characteristics of failed angioplasty that enhanced the information provided by coronary angiography and assisted in the selection of the bailout therapy.

Vessel reconstruction in total coronary occlusions with a long subintimal wire pathway: use of multiple stents under guidance of intravascular ultrasound.

A frequent cause of failure of the recanalization of a total coronary occlusion is a subintimal pathway of the guide wire. Three cases of occluded right coronary arteries are presented in which a distal reentry into the true vessel lumen was achieved. Intravascular ultrasound was used to locate the exit and reentry of the guide wire, and to plan the position of multiple stents for the coverage of this subintimal pathway. In all cases antegrade flow to the distal coronary bed was restored.

Morphology of chronic coronary occlusions and response to interventional therapy--a study by intracoronary ultrasound.

OBJECTIVES: Balloon angioplasty of chronic coronary occlusions has a low procedural success and a high recurrence rate. Better tomographic insights into the lesion morphology may improve the interventional strategy and results. METHODS: Intracoronary ultrasound was used during the recanalizaton procedure of 45 chronic coronary occlusions (2 weeks to 14 months; average 3.4 months) to determine the lesion morphology and to assess the angioplasty result. The luminal area and the plaque burden were measured proximal and distal to the occlusion, and within the occlusion. The ultrasonographic characteristics of the occlusive lesions were compared to 45 nonocclusive lesions of age-matched patients with stable angina pectoris. RESULTS: Occlusive lesions were more often echodense as compared to nonocclusive lesions (35% vs. 20%; p = 0.10). In chronic occlusions a multi-layered plaque morphology was observed in 22%, and this morphology was not found in nonocclusive lesions. Angiographic characteristics were not related to the ultrasonographic morphology of the lesion. Despite similar vessel areas in occlusive and nonocclusive lesions, the balloon size selected according to the angiographic image was underestimated in occlusive lesions. Based on the quantitative ultrasound measurement the balloon size was increased from 2.6 +/- 0.3 mm to 3.3 +/- 0.5 mm in 53% of the lesions. This resulted in an increase of the luminal area from 3.51 +/- 0.92 to 5.08 +/- 1.43 mm2 (p < 0.001). The acute recoil after balloon angioplasty was similar (34 +/- 18%) in hypodense and echodense plaques, but was significantly higher in lesions with a multi-layered plaque morphology (49 +/- 22%; p < 0.05). In 19 patients with severe dissections or extreme acute recoil (residual stenosis > 50%) the use of a stent increased the luminal area from 3.94 +/- 0.81 to 7.51 +/- 1.71 mm2 (p < 0.001). CONCLUSIONS: Intracoronary ultrasound demonstrated a multi-layered plaque morphology in one fourth of the chronic occlusions. This type of plaque was associated with a significant acute recoil. The presence of diffuse atherosclerosis in neighbouring segments of chronic coronary occlusions leads to underestimation of the balloon size. Quantitative assessment by intracoronary ultrasound helped to optimize the balloon size leading to a significant luminal area gain. The detection of excessive acute recoil should be considered an indication for stent deployment.

Sonographic and Angiographic Features of Intramural Hematoma as a Cause of Failed Coronary Angioplasty.

A characteristic feature of a specific type of dissection after PTCA was observed in seven cases with severe obstruction of the vessel segments proximal and/or distal to the dilatation site. The underlying cause in the absence of an angiographically visible dissection flap was revealed by intravascular ultrasound. The obstruction was caused by an extraluminal obstruction by an echodense mass increasing in size with time, and without a dissection flap even as assessed by intravascular ultrasound. This phenomenon was interpreted as an intramural hematoma extending between media and adventitial border of the vessel. In six of seven cases, bailout therapy by either stenting or redilatation and creation of a typical dissection flap relieved the vessel obstruction. In one case of extension of the hematoma into the left main coronary artery, CABG was performed. Intravascular ultrasound provided a conclusive insight into the underlying morphology of failed PTCA in cases without angiographic features of a dissection. It helped in deciding and controlling the bailout strategy, which was the stenting of the entry into the hematoma at the initial dilatation site.

Behavioral risk factors of Chippewa Indians living on Wisconsin reservations.

Behavioral risk factors and chronic disease death rates vary markedly among the numerous American Indian tribes. Local data on prevalence of risk factors are important in determining effective community-based interventions. The authors conducted an in-person survey to ascertain the prevalence of behavioral risk factors among members of the Chippewa tribe living on reservations in Wisconsin. A total of 465 Chippewa adults were randomly selected from tribal registries and invited to participate in the study. Of these, 175 (38 percent) participated. To characterize nonrespondents, 75 nonrespondents were randomly selected and aggressively followed up. The authors compared their results with data from the 1989 Wisconsin Behavioral Risk Factor Surveillance System. Chippewa respondents reported high levels of obesity and tobacco use. No significant differences existed between the original survey and followback of nonrespondents. Compared with respondents who had telephones, those without telephones were significantly more likely to be unemployed, to be a current smoker or drinker, and to report nonuse of seatbelts. Compared with the general Wisconsin population, Chippewa adults appear to have higher prevalences of several chronic disease and injury risk factors. The original survey methodology, despite the low response rate, appeared to give a more accurate (less biased) estimate of risk factor prevalences than would have been achieved by a telephone survey.

Fish consumption patterns and blood mercury levels in Wisconsin Chippewa Indians.

Methylmercury is a known neurotoxin at high blood levels (> 400 micrograms/l) and is thought to cause neurologic symptoms at substantially lower levels in susceptible adults and infants. Given that levels of methylmercury in fish in northern Wisconsin lakes can be high (> 1 ppm, FDA standard) and Chippewa Indians take large amounts of fish from these lakes, the extent of their exposure to methylmercury was investigated. Using tribal-maintained registries, 465 Chippewa adults living on reservation were selected randomly and were invited to participate; 175 (38%) participated in the study. In an effort to characterize nonrespondents, 75 nonrespondents were selected randomly and were followed up aggressively. An additional 152 volunteers who were selected nonrandomly also participated in the study. Subjects completed a questionnaire about fish consumption patterns and had blood drawn for mercury determination. Sixty-four persons (20%) had blood mercury levels in excess of 5 micrograms/l (i.e., upper limit of normal in nonexposed populations); the highest level found was 33 micrograms/l. Fish consumption was higher in males and the unemployed. Blood mercury levels were highly associated with recent walleye consumption (p = .001). Methylmercury levels in some Wisconsin Chippewa were found to be elevated, but were below the levels associated with adverse health effects. We recommend a continuation of efforts to limit exposures in this high-risk population.

Increased expression of heat shock protein, transferrin, and beta 2-microglobulin in astrocytes during scrapie.

Scrapie is a slow infection and neurodegenerative disease of animals characterized pathologically by formation of amyloid, astrocytosis, vacuolation and injury and death of neurons. Our previous studies of scrapie point to: (i) a critical role for the astrocyte in responding to, and perhaps inadvertently contributing to, the neuropathological manifestations of infection; and (ii), the hypothesis that the astrocyte executes a programed response to neurological injury analogous to the stress response. The expression of genes encoding transferrin and beta 2-microglobulin has been shown to increase during scrapie and we sought in the studies reported here to determine whether this modulated expression would map to astrocytes. We also looked for changes in a heat shock protein that is induced in the stress response. We show that transferrin, beta 2-microglobulin and heat shock 72 kD protein all increase in astrocytes in the course of infection. We speculate in the discussion on the possible functions of these and other proteins in neurodegenerative processes and why these functions so frequently reside in the astrocyte.

Neuropathological changes in scrapie and Alzheimer's disease are associated with increased expression of apolipoprotein E and cathepsin D in astrocytes.

With the rationale that the neuropathological similarities between scrapie and Alzheimer's disease reflect convergent pathological mechanisms involving altered gene expression, we set out to identify molecular events involved in both processes, using scrapie as a model to study the time course of these changes. We differentially screened a cDNA library constructed from scrapie-infected mice to identify mRNAs that increase or decrease during disease and discovered in this way two mRNAs that are increased in scrapie and Alzheimer's disease. These mRNAs were subsequently shown by sequence analysis to encode apolipoprotein E and cathepsin D (EC 3.4.23.5). Using in situ hybridization and immunocytochemistry to define the cellular and anatomic pathology of altered gene expression, we found that in both diseases the increase in apolipoprotein E and cathepsin D mRNAs and proteins occurred in activated astrocytes. In scrapie, the increase in gene expression occurred soon after the amyloid-forming abnormal isoform of the prion protein has been shown to accumulate in astrocytes. In Alzheimer's disease, the increased expression of cathepsin D also occurred in association with beta-amyloid. These studies reveal some of the molecular antecedents of neuropathological changes in scrapie and Alzheimer's disease and accord new prominence to the role of astrocytes in neurodegenerative conditions.