Influence of lectins, hexoses, and neuraminidase on the association of purified elementary bodies of Chlamydia trachomatis UW-31 with HeLa cells.

Using highly purified elementary bodies of Chlamydia trachomatis UW-31 (serotype K), we found that HeLa 229 monolayer cultures bound more 32P-labeled chlamydiae after pretreatment with the lectin wheat germ agglutinin. The lectin, on the other hand, inhibited competitively when chlamydial association was assayed in the presence of polycations. The two effects of wheat germ agglutinin were abolished when N-acetylneuraminic acid (NeuNAc)- or N-acetylglucosamine (GlcNAc)-preincubated wheat germ agglutinin was used. Brief exposure of HeLa cells to neuraminidase abolished the ability to bind the elementary bodies, whether or not polycations were present. Furthermore, at 5 degrees C but not at 37 degrees C, NeuNAc, GlcNAc and N-acetylgalactosamine inhibited chlamydial association only in the absence of the polycation DEAE-dextran. The results suggest that NeuNAc residues on the plasma membrane are the principal, but not the only, receptors for this strain of C. trachomatis.     

Genetic recombination of bacteriophage T7 DNA in Vitro IV. Asymmetry of recombination frequencies caused by polarity of DNA packaging in Vitro

Recombination frequencies of phage T7 DNA measured using an in vitro assay for genetic recombination and DNA packaging (Sadowski and Vetter, 1976) show a marked asymmetry. This asymmetry was shown to be caused by polar packaging of the DNA during the reaction rather than asymmetry of recombination itself. Recombinant T7 DNA, formed during a physical assay for in vitro T7 recombination (Vlachopoulou and Sadowski, 1977), was shown to be genetically recombinant using a highly efficient assay for packaging of T7 DNA in the absence of recombination.     

“Porcupine” polymers and double-star polymers

Anionic block copolymerization of styrene and divinylbenzene is known to lead to the formation of star-shaped macromolecules. This “arm-first” method has been widely used and studied. The present paper is devoted to two special aspects of this method: The first is concerned with the efficiency of the protection exerted by the arms on the crosslinked core, preventing gelation of the reaction medium. A number of “porcupine” polymers involving bulky cores, fitted with a large number of arms, were synthesized and characterized. The second deals with the possibility of using the “living” carbanionic sites present in the cores, either for purpose of functionalization, or to grow new branches from the core. The presence of remaining unsaturations in the core was evidenced. This is a drawback, since the possibility for “transverse” bonds to be formed results in couplings, inducing gelation.     

Effect of age on humoral immunity, selection of the B-cell repertoire and B-cell development

Summary: The age-associated changes in humoral Immunity affect the quality more than the quality of the antibody response. Changes in the quality of the antibody response with age include shifts in antibody specificities from foreign to autoantigens. in antibody isotypes from IgG to IgM, in antibody affinities from HIGH to low and in the antibody idiotypic repertoire. These changes can be traced to an impaired capacity of T cells to facilitate: (a) the maturation of B cells respect to isotype and affinity maturation in the periphery and (b) the development of a diverse B-cell repertoire from precursors within the bone marrow. In contrast, there is no evidence that the amount of immunoglobulin produced before or after immunization diminishes with age. Nonetheless, the impaired responses of the elderly to most vaccines and the greater susceptibility of the elderly to infections has fostered a view that immune senescence leads to a state of immune deficiency. However, it is more precise to describe immune senescence as leading to a state of immune dysregulation. The dysregulation of the humoral immunity is manifested by a shift from adaptive humoral Immunity, characterized by the production of a highly specific, high-affinity, IgG antibody response to foreign antigens, to a process of natural antibody-mediated immunity, dominated by low-affinity, polyreactive, IgM antibodies which react with autoantigens, Age-associated T-cell impairments appear to be the basis for the shift from adaptive to natural humoral immunity and their reversal should permit the restoration of an adaptive antibody response in the elderly.     

Differentiation of Normal and Malignant Human Squamous Epithelium in Vivo and in Vitro: A Morphologic Study

We would like to thank John Ellis for expert photographic assistance, Mervin Jones and Linda Lovell for expert animal husbandry, and Dr. Dorothy Easty for establishing the cell line LICR-LON-HN-5, without which this study would not have been possible.

We report a light microscopic and ultrastructural analysis of the comparative degrees of differentiation seen in keratinocytes derived from the tongue and epidermis with those of a well-differentiated human squamous carcinoma cell line (LICR-LON-HN5). When growing on plastic substrates, all cultures had a similar morphology, with multilayering and the production of cornified envelopes. When cultured on collagen gels the structure was more organized, with keratohyalin granules and keratin whorl formation in both the normal and the malignant cultures. Normal keratinocytes injected into athymic mice produced epidermal cysts, while cells from the cell line produced well-differentiated squamous cell carcinomas, which were partially solid and partially cystic. The tumor was well organized, with identifiable basal cells, spin-ous cells, keratohyalin granules, and a prominent basal lamina at the stromal/epithelial interface. This model is to be developed for comparative studies between normal and malignant cells, with particular reference to basement membrane production and to investigations of the relative importance of extrinsic and intrinsic factors in the control of squamous differentiation.     

Loss of imprinting of IGF2 and H19 in osteosarcoma is accompanied by reciprocal methylation changes of a CTCF-binding site

The adjacent insulin-like growth factor 2 (IGF2) and H19 genes are imprinted in most normal human tissues, but imprinting is often lost in tumors. The mechanisms involved in maintenance of imprinting (MOI) and loss of imprinting (LOI) are unresolved. We show here that osteosarcoma (OS) tumors with IGF2/H19 MOI exhibit allele-specific differential methylation of a CTCF-binding site upstream of H19. LOI of IGF2 or H19 in OS occurs in a mutually exclusive manner, and occurs with monoallelic expression of the other gene. Bisulfite sequencing reveals IGF2 LOI occurs with biallelic CpG methylation of the CTCF-binding site, while H19 LOI occurs with biallelic hypomethylation of this site. Our data demonstrate that IGF2 LOI and H19 LOI are accompanied by reciprocal methylation changes at a critical CTCF-binding site. We propose a model in which incomplete gain or loss of methylation at this CTCF-binding site during tumorigenesis explains the complex and often conflicting expression patterns of IGF2 and H19 in tumors.     

Expression of alpha-methylacyl-CoA racemase (P504s) in various malignant neoplasms and normal tissues: astudy of 761 cases

Alpha-methylacyl CoA racemase (AMACR), also known as P504S, plays an important role in peroxisomal beta-oxidation of branched-chain fatty acids. It has recently been shown that AMACR is highly expressed in prostate cancer and that it may be an important diagnostic marker for prostate carcinoma. However, little is known about expression of AMACR in normal tissues and other malignant tumors. In this study, we investigated expression of AMACR in 539 malignant tumors and 222 normal human tissues of various types by immunohistochemical analysis. mRNA levels of AMACR in normal organs and in selected tumors were assessed by real time PCR. In normal tissue, high expression of AMACR mRNA was identified in liver, kidney and salivary gland, while AMACR protein was detected in liver (hepatocytes), kidney (tubular epithelial cells), lung (only bronchial epithelial cells), and gallbladder (only mucosal epithelial cells). High expression of AMACR mRNA was found in prostate, liver, and kidney cancers but rarely in stomach and bladder cancers. A high percent of adenocarcinomas arising from these organs express AMACR, including 17 of 21 (81%) of hepatocellular carcinomas and 18 of 24 (75%) of renal cell carcinomas. In addition, carcinomas arising from tissues normally not expressing AMACR were also positive for the antigen, including 17 of 18 (94%) prostate carcinomas, 9 of 29 (31%) of urothelial carcinomas, and 4 of 15 (27%) of gastric adenocarcinomas. Two hundred and fifty cases of adenocarcinomas from lung, breast, pancreas, bile duct, adrenal gland, salivary gland, ovary, thyroid and endometrium were negative or rarely positive for AMACR. Neuroendocrine carcinomas rarely expressed AMACR. Melanomas, squamous cell carcinomas, basal cell carcinomas, soft tissue tumors (including epithelioid sarcomas and synovial sarcoma), thymomas, and germ cell tumors were negative for AMACR. Our data provide important baseline information for using AMACR in clinical practice and also are valuable in furthering understanding of the pathogenic role of AMACR in malignant neoplasms.     

Sociotropic cognition moderates stress-induced cardiovascular responsiveness in women through effects on total peripheral resistance, but not cardiac output.

Although cardiovascular disease (CVD) remains the leading cause of mortality in women, few studies have examined the role of psychosocial factors in its development. This study examined the moderating effects of sociotropic cognition (SC), a need for social acceptance and approval, on psychosocial stress-induced cardiovascular responsiveness (CVR) and affect reactivity in women. Sixty-eight normotensive, college-aged females were randomly assigned to a low or high social threat condition. Measures of systolic, diastolic and mean arterial blood pressures (SBP, DBP and MAP, respectively), heart rate (HR), cardiac output (CO), total peripheral resistance (TPR) and negative affect were collected during rest, and under conditions of high vs. low interpersonal threat. A two-step hierarchical regression analysis was performed to predict all response variables (BPs, HR, CO, TPR and affect). Increases in SBP, DBP, MAP, TPR and negative affect were greater in the high threat than low threat condition. Changes in SBP, MAP and TPR positively covaried with SC under conditions of high interpersonal threat, but showed no significant covariation in the low threat condition. The data suggest that an excessive need for social acceptance may contribute to rises in BP through an increase in TPR, but not CO under conditions of high social threat.     

The therapeutic alliance in cognitive-behavioral treatment of children referred for oppositional, aggressive, and antisocial behavior

The authors examined the therapeutic alliance in evidence-based treatment for children (N = 185, 47 girls, 138 boys; ages 3-14 years) referred clinically for oppositional, aggressive, and antisocial behavior. Different alliances (child-therapist, parent-therapist) were assessed from each participant's perspective at 2 points over the course of treatment. As predicted, both child-therapist and parent-therapist alliances related to therapeutic change, family experience of barriers to participation in treatment, and treatment acceptability. Greater alliance was associated with greater therapeutic change, fewer perceived barriers, and greater treatment acceptability. The findings could not be attributed to the influence of socioeconomic disadvantage, parent psychopathology and stress, and child dysfunction or to rater effects (common rater variance in the predictors and criteria).     

Sensitivity of Fluorochrome Microscopy for Detection of Mycobacterium tuberculosis versus Nontuberculous Mycobacteria

The results for 6,532 consecutive mycobacterial respiratory specimens collected from 1,040 patients from 1993 to 1995 in a Texas hospital were studied to determine the sensitivity of fluorescence microscopy for detection of Mycobacterium tuberculosis and nontuberculous mycobacteria (NTM). Smears were positive for acid-fast bacilli (AFB) in 63% (677 of 1,082) of specimens growing M. tuberculosis and 56% (638 of 1,148) of specimens growing the four most common species of NTM. Smear positivity by species was 58% (446 of 776) for M. avium complex, 51% (154 of 300) for rapidly growing mycobacteria (98% were M. abscessus), 78% (29 of 37) for M. kansasii, and 26% (9 of 35) for M. gordonae. Definite or probable disease by clinical criteria was present in 79% of patients with M. avium complex, 93% of patients with rapidly growing mycobacteria, 100% of patients with M. kansasii, and 0% of patients with M. gordonae. Patients with M. avium complex had a low incidence of AIDS (7%), and approximately 50% of non-AIDS patients had upper-lobe cavitary disease and 50% had nodular bronchiectasis. Only 23 of 6,532 (0.35%) of AFB smears were positive with a negative culture excluding patients on therapy for established mycobacterial disease. These studies suggest that NTM are as likely as M. tuberculosis to be detected by fluorescent microscopy in specimens from patients from areas endemic for NTM lung disease and at low risk for AIDS.