Characterization of the unusual basic proteins of cricket spermatid nuclei on the basis of their molecular weights and amino acid compositions

Typical somatic cell type histones are lost from the nucleus during late spermiogenesis in the house cricket; they are replaced by unusual basic proteins specific to the spermatid. We wish to characterize these proteins because they appear to determine the unusual chromatin structures of the spermatid. Molecular weights of the unusual basic proteins were estimated by chromatographing them on Bio-Gel A 0.5 M agarose columns eluted with 6 M guanidine hydrochloride. Two proteins named TH1 and TH2 have molecular weights in the range spanned by the somatic histones. The molecular weight of TH1 is 17 500 and that of TH2 is 15 500. Three additional spermatid proteins were also analyzed by molecular weight determination. They are called here protamines A, B and C, and they have molecular weights in the range typical of protamines. That of A is 6200, of B is 5500 and of C is 3800. They span the range from the large protamines typical of mammalian sperm to the small protamines of salmonid fish. The molecular weights of the TH proteins were also examined by electrophoresis on SDS-polyacrylamide gels. Amino acid compositions determined for TH1 and TH2 show that both are basic proteins rich in arginine relative to lysine. Their compositions are histone-like, but they appear to be distinct histone types rather than variant forms of the somatic histones.     

Reduced cerebrospinal fluid levels of immunoreactive pro-opiomelanocortin related peptides (including beta-endorphin) in anorexia nervosa

The discovery that the endogenous opioid peptides contribute to the modulation of appetitive behavior and neuroendocrine function has raised questions as to whether disturbances of opioids contributes to the pathophysiology of eating disorders. To assess central nervous system (CNS) beta-endorphin in patients with anorexia nervosa we measured cerebrospinal fluid (CSF) beta-endorphin concentrations before, and at intervals after weight correction. In addition, we measured three sister peptides (beta-lipotropin, adrenocorticotropic hormone (ACTH), and the N-terminal fragment) derived from the same precursor molecule, pro-opiomelanocortin (POMC) to determine whether possible disturbances might extend to sister peptides. Underweight anorectics (58 +/- 5% of average body weight (ABW), n = 10) had significantly lower CSF concentrations of all 4 peptides compared to healthy controls (102 +/- 10% ABW, n = 11). CSF concentrations of all 4 POMC-related peptides were found to be significantly increased when the same anorectics were restudied 4 to 6 weeks after weight gain (83 +/- 4% ABW). After weight gain, levels of CSF beta-endorphin, beta-lipotropin, and ACTH were similar to controls, whereas levels of CSF N-POMC remained significantly less than controls. Another group of women, previously underweight with anorexia nervosa, but weight-restored (93 +/- 11% ABW, n = 12) for greater than 1 year had CSF concentrations of all 4 POMC-related peptides that were similar to controls. We conclude that underweight anorectics have state-associated disturbances of CNS beta-endorphin as well as other POMC-related peptides. These abnormalities are part of the neurobiological syndrome of anorexia nervosa and may contribute to the characteristic alterations in behavior and neuroendocrine function.     

Enzyme-linked immunosorbent assays for detecting antibodies to Shiga-like toxin I,Shiga-like toxin II,andEscherichia coli O157:H7 lipopolysaccharide in human serum

Shiga-like toxin-producingEscherichia coli O157:H7 are important causes of bloody diarrhea and hemolytic uremic syndrome. To facilitate the epidemiologic study of these organisms, we developed enzyme-linked immunosorbent assays (ELISAs) for antibodies to Shiga-like toxin I (SLT I), Shiga-like toxin II (SLT II), andE. coli O157 lipopolysaccharide (LPS). We tested serum samples from 83 patients in two outbreaks ofE. coli O157:H7 diarrhea and from 66 well persons. Forty-three patients (52%) had at least one serum sample positive for anti-O157 LPS antibodies; among 26 culture-confirmed patients, 24 (92%) had at least one positive serum sample. Two (3%) of 66 control sera had positive anti-O157 LPS titers. ELISA results for SLT I and II were compared with those of HeLa cell cytotoxicity neutralization assays on both patient and control sera. Neutralization assays detected anti-SLT I antibodies in at least one serum sample from each of 17 (20%) patients and 7 (10.6%) controls, while 16 (19%) patients and 7 controls had positive titers by anti-SLT I ELISA. Although all serum samples, including control sera, showed nonspecific neutralization of SLT II, no antibody titers to SLT II were detected by either neutralization or ELISA. These results indicate that ELISAs for SLT I and SLT II antibodies are comparable to HeLa cell cytotoxicity neutralization assays. Both the ELISAs and neutralization assays are insensitive in detecting infected patients. However, the ELISA for antibodies toE. coli O157 LPS is both sensitive and specific, and may be more useful than assays for antitoxic antibodies in detecting persons withE. coli O157:H7 infection.     

Genetic basis for altered pathogenesis of an immune-selected antigenic variant of reovirus type 3 (Dearing).

In this paper we provide a step by step comparison of the pathogenesis of murine infection caused by reovirus type 3 (Dearing) and an antigenic variant (K) selected by its resistance to neutralization with a monoclonal antibody (G5) directed against the T3 hemagglutinin. To show that specific changes in the biologic properties of variant K were due to mutation in the S1 double-stranded RNA segment (gene), which encodes the viral hemagglutinin, we generated a reassortant virus ("1 HA K") containing the variant K S1 gene and compared its properties to variant K and to a reassortant ("1 HA 3") containing the T3 (Dearing) S1 gene. These studies, in conjunction with our previous nucleotide sequence analysis of the S1 genes of variant K and T3 (Dearing) [R. Bassel-Duby, A. Jayasuriya, D. Chatterjee, N. Sonenberg, J. V. Maizel, Jr., and B. N. Fields, Nature (London) 315:421-423, 1985; R. Bassel-Duby, D. R. Spriggs, K. L. Tyler, and B. N. Fields, submitted for publication], indicate that a single amino acid change in the T3 hemagglutinin can alter viral growth and tropism within the central nervous system without affecting either its primary replication in the intestine or its pattern of spread to or within the central nervous system.     

Stimulation of creatine kinase BB activity by parathyroid hormone and by prostaglandin E2 in cultured bone cells.

Bone cells in culture responded to parathyroid hormone (PTH) and prostaglandin E2 (PGE2) by a 2-fold increase in creatine kinase (CK) activity. Combined treatment resulted in a higher response than with PTH alone. Calcitonin (CT) failed to stimulate CK activity, did not affect the response of CK to PTH, but inhibited slightly the increase in CK activity by PGE2. Bone-cell cultures grown in low [Ca2+] (0.125 mM), enriched in PTH-responsive osteoblast-like cells, responded to PTH, but not to PGE2 or CT, by increased CK activity. In both normal and low-[Ca2+] cultures, 8-bromo cyclic AMP did not affect CK activity, nor did it change the response of the cells to PTH, PGE2 or CT. The increase in CK activity was time- and dose-dependent and inhibited both by cycloheximide and by actinomycin D. The isoenzyme of CK stimulated was the CKBB form, the isoenzyme induced by other hormones. This appears to be the first report of the stimulation of CK activity by a polypeptide hormone or a prostaglandin. We suggest that stimulation of CKBB can serve as a marker for the action of a variety of hormones and growth promoters.     

In Vitro Antimicrobial Activity and Human Pharmacology of Cephalexin, a New Orally Absorbed Cephalosporin C Antibiotic

Concentrations of cephalexin (an orally absorbed derivative of cephalosporin C) in serum and urine were determined in normal volunteers and patients. The in vitro antibacterial activity was also studied. All strains of group A β-hemolytic streptococci and Diplococcus pneumoniae were inhibited by 3.1 μg/ml. Of the Staphylococcus aureus strains, 88% were inhibited by 6.3 μg/ml, and 12.5 μg/ml was inhibitory for all S. aureus, 80% of Escherichia coli, 72% of Klebsiella-Aerobacter, and 56% of Proteus mirabilis strains. About 90 to 96% of E. coli, Klebsiella Aerobacter, and P. mirabilis strains were inhibited by 25 μg of cephalexin per ml. Pseudomonas and indole-positive Proteus strains proved to be quite resistant to cephalexin. Cephalexin was well absorbed after oral administration. A peak serum concentration of cephalexin of at least 5 μg/ml was achieved in each volunteer with 250 and 500-mg doses. A mean peak serum concentration of 7.7 μg/ml was achieved with 250-mg doses; 12.3μg/ml was achieved with 500-mg doses of antibiotic. Food did not interfere with absorption. Probenecid enhanced both the peak serum concentration and the duration of antibiotic activity in the serum. Over 90% of the administered dose was excreted in the urine within 6 hr. The mean peak serum concentration of cephalexin after an oral dose of 500 mg was adequate to inhibit all group A streptococci, D. pneumoniae, and S. aureus, 85% of E. coli, and about 40 to 75% of Klebsiella-Aerobacter and P. mirabilis strains. Levels of cephalexin in urine were adequate to inhibit over 90% of E. coli, and P. mirabilis and 80 to 96% of Klebsiella-Aerobacter strains.     

The Role of Calcium Ions in the Acceleration of Resting Muscle Glycolysis by Extracellular Potassium

The activation of the glycolysis of resting muscle by increased extracellular potassium is dependent upon the simultaneous presence of calcium, but not of sodium ions. This regulation of metabolism by a membrane characteristic seems to act upon an early link in the glycolytic enzyme chain.     

Cerebrospinal fluid gradients of acetylcholinesterase and butyrylcholinesterase activity in healthy aging

Acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) activities were measured in 13 sequential 2 ml aliquots of cerebrospinal fluid (CSF) obtained by lumbar puncture from 7 young and 7 elderly healthy normal subjects. The slopes of the rostrocaudal gradients of AChE and BChE were calculated and compared to those of total protein concentration and the major dopaminergic metabolite homovanillic acid (HVA), for which a pronounced rostrocaudal gradient (with highest concentrations of HVA in more rostral CSF) is consistent with HVA originating primarily from the brain. AChE activity was higher in more caudal fractions of young, but not elderly subjects and there was a significant difference between the mean AChE gradient slopes in the young and old groups. These results suggest that the spinal cord makes an important contribution to AChE activity in lumbar CSF. Furthermore, the absence of a negative AChE gradient in elderly subjects may be the result of a greater rate of entry of cerebral AChE into CSF, possibly as a consequence of an increased ventricular surface area and shorter diffusion distances in atrophic elderly brains. In contrast to AChE, BChE activity and total protein concentrations were higher in more caudal CSF fractions of not only young but also old subjects. In addition, there was a significant correlation between the gradient slopes of BChE activity and total protein concentrations, suggesting that the majority of BChE activity in lumbar CSF derives from the same source as the majority of total protein, namely plasma. The diffuse (i.e. brain and spinal cord) origin of AChE in lumbar CSF would explain the relatively modest changes in lumbar CSF AChE activity in diseases involving certain central cholinergic systems, most notably Alzheimer's disease.