Edward C. Franklin

Mixed cryoglobulinemia: Clinical aspects and long-term follow-up of 40 patients☆

The clinical course of 40 patients with significant quantities of mixed cryoglobulins, but without lymphoproliferative, collagen-vascular or chronic infectious diseases, is presented. These cases comprise 51.3 percent of all mixed and 31.7 percent of all types of cryoglobulins evaluated by us over the period 1960--1978. A characteristic clinical syndrome, consisting of recurrent palpable purpura (100 percent), polyarthralgias (72.5 percent) and renal disease (55 percent), was seen. Biopsy specimens of skin lesions showed cutaneous vasculitis, and half had immune reactants in vessel walls. Seventy percent of patients had evidence of hepatic dysfunction, often subclinical, and more than 60 percent of those tested had serologic evidence of prior infection with hepatitis B virus. Hepatic lesions ranged from minimal triaditis to chronic active hepatitis and/or cirrhosis. All 22 patients in whom clinical renal disease developed had significant proteinuria; 63.6 percent had diastolic hypertension, 77.3 percent edema, 45.5 percent renal failure and 22.7 percent were nephrotic. Glomerular disease associated with deposition of immunoglobulin G, immunoglobulin M and complement, often with coexistent renal arteritis, was confirmed pathologically in 15 cases. All cryoglobulins had rheumatoid factor activity and consisted of IgM and polyclonal IgG; five also contained IgA. Thirteen had a monoclonal IgM kappa component. Serum protein electrophoresis was unremarkable or showed diffuse hyperglobulinemia. Striking depression of early complement components was noted but did not correlate well with the cryoprotein concentration, renal involvement or clinical course. Follow-up for periods up to 21 years from onset of symptoms revealed that renal involvement has a deleterious effect on prognosis. Postmorten examinations of nine patients demonstrated widespread vasculitis in addition to renal involvement. Preterminal infection was found in eight.

The characterization of soluble amyloid prepared in water

Amyloid was extracted from the spleen of a patient with primary amyloidosis by homogenizing it at high speed with water after preliminary treatments, first to remove proteins soluble in saline, and then to remove salts. The extracts containing amyloid appeared to be clear at concentrations up to 6 mg/ml of protein. The material gave little sediment on being centrifuged up to 20,000 g for 1 hr, but the protein was sedimented at 100,000 g in 1 hr. The amyloid could be precipitated from the extracts by addition of NaCl to 0.0075 mole/liter or of CaCl(2) to 0.0025 mole/liter. The protein-bound Congo red formed a red precipitate and this property was used to estimate recovery and purity of amyloid during extraction. On electronmicroscopy the isolated amyloid proved to be morphologically pure. It existed either as single filaments measuring 60-80 A in diameter or as large aggregates of these filaments.Freshly isolated amyloid in water sedimented as a single homogeneous peak with an s degrees (20,[unk]) of about 45-50S. On standing, the solution became cloudy and more rapidly sedimenting components appeared. On electrophoresis the material migrated as a homogeneous peak towards the anode. The protein had an amino acid composition different from that of all known serum proteins. It was rich in acidic amino acids and had little cysteine and methionine and no hydroxyproline. The total content of carbohydrate was less than 2%.

Cryoglobulinemia—A study of twenty-nine patients: I. IgG and IgM cryoglobulins and factors affecting cryoprecipitability☆

Abstract A study of twenty-nine patients with cryoglobulinemia revealed the presence of G immunoglobulins in eight (generally found in patients with multiple myeloma or idiopathic cryoglobulinemia), macroglobulins in nine and unusual cryoglobulins with rheumatoid factor activity in twelve. Among the latter patients, a polymer of IgG was observed in a patient with rheumatoid arthritis and mixed cryoglobulins composed of IgM and IgG globulins were found in the remaining eleven. Factors affecting cryoprecipitability and the stability of the cryoglobulins were studied. Cryoprecipitation was inhibited by extremes of pH and ionic strength, and by 2M urea. Sulfhydryl reagents reversibly destroyed IgM cryoglobulins, but had little if any effect on IgG cryoglobulins. No specific structure could be related to cold insolubility. Symptoms possibly due to the presence of cryoglobulins appeared to be related only to some extent to their concentration, but more importantly to the physicochemical properties of these proteins and the temperature at which precipitation occurred.

Proteolytic enzymes from the mouse submaxillary gland. Specificity restricted to arginine residues.

Abstract The mouse submaxillary proteases (A + D), the isolation and properties of which were previously described by us, hydrolyze only arginyl bonds in proteins. Formol titrations and peptide mapping on digests of polyarginine, polylysine, lysozyme, histone, and insulin suggested this specificity. Amino acid compositions of peptides from lysozyme and insulin showed that most but not all arginyl bonds were hydrolyzed but that no lysyl bonds were split. The proteases should be useful in the sequencing of proteins.

Complement fixation by antibody fragments.

Rabbit antibodies (7S) degraded by papain into univalent 3.5S fragments fail to fix complement when they combine, but do not precipitate, with the homologous antigens. Divalent 5S fragments obtained by pepsin digestion (composed of fragment I linked to II, but lacking fragment III) also fail to fix complement although they precipitate with homologous antigens. The amount of specific precipitate formed by the 5S antibody fragment is not increased by exposure to complement

Staphylococcal Protein A and Human IgG Subclasses and Allotypes

Staphylocoecal protein A binds molecules belonging to the IgGl, IgG2, and IgG4 sub‐classes. IgG3 proteins generally do not bind, except for those coded by the two γ3 alleles, which are G3m(u−): G3m(b0,b3,b5,s,v) and G3m(b0,b3,b5,s,t,v). G3m(u) is located in the CH2 domain. The difference between G3m(u−) and G3m(u+) IgG3 proteins correlates with the sequence at position 339 in the CH2 domain—Ala and Thr respectively. There is another structural difference in the CH3 domain which correlates with protein A binding and non‐binding: all IgG proteins that bind protein A have His at position 435, whereas those that do not, have Arg at that position.

Two Types of γ-Globulin Differing in Carbohydrate Content

Summary 1. The heavy components (19 S) have been highly concentrated from normal human serum by repeated cycling in the preparative ultracentrifuge. Electrophoresis of such preparations has permitted isolation of a high molecular weight γ-globulin fraction. 2. Analyses of this material indicate a high carbohydrate content with hexose sugars and sialic acid present at approximately 5 times the concentration found for the main 7 S γ-globulin. The hexosamine-hexose ratio is approximately 0.5. 3. Markedly reduced levels were found in the sera of 4 patients with agammaglobulinemia. The possibility that certain antibodies fall into the 19 S fraction and are similarly rich in carbohydrate is discussed.

Amyloid-related serum component (SAA)--studies in acute infections, medullary thyroid carcinoma, and postsurgery.

Abstract Longitudinal studies of serum SAA (amyloid-related serum component) levels were done on a group of 17 patients hospitalized for acute bacterial infections, and the results were compared to another group of 24 patients undergoing elective surgical procedures. In the former, striking initial elevations were seen, resolving rapidly with therapy and clinical status; failure of response in two patients could be correlated with ongoing or recurrent infection. By contrast, the latter group manifested little variation in SAA level. Of all the acute-phase parameters tested, only the WBC (white blood cell count) could be related to variations in SAA. Levels obtained in patients and relatives of patients with the familial form of medullary carcinoma of the thyroid were generally normal. The possible significance of this finding is discussed in view of theories claiming the amyloid associated with this entity to be light-chain-related, “A” protein, or prohormone in nature. SAA was found to be present at low levels in CSF (cerebrospinal fluid), though no correlation to age could be made at the level of resolution of the radioimmunoassay. Simultaneous serum levels ranged up to 30 times the CSF levels. A small group of patients with elevated CSF levels were also noted to have acute infections, some without evidence of meningeal irritation.

Current concepts of amyloid.

Publisher Summary This chapter discusses the structure, chemistry, immunology, cellular origin, metabolism, and possibly the pathogenesis of amyloid. The simplest and most widely used classification of amyloidosis is based on the four major categories introduced by Reimann. Primary amyloid occurs with no known antecedent or coexistent disease and usually involves mesodermal tissues, such as smooth or skeletal muscle, or the cardiovascular system. Generally there is variability in staining of the deposits with Congo red, iodine, and metachromatic dyes. Secondary amyloid is usually associated with chronic diseases, such as infections, neoplasms, neurological disorders, or connective tissue diseases—especially rheumatoid arthritis. It generally involves spleen, liver, kidney, intestines, and adrenals and tends to have reproducible and characteristic staining properties with the above mentioned dyes. Amyloid associated with myeloma tends to resemble the primary type but is invariably associated with a neoplasm involving plasma cells or lymphocytes—such as multiple myeloma, macroglobulinemia, or heavy chain disease. Tumor-forming amyloid is characterized by the small masses of amyloid in the skin, eye, bladder, urethra, respiratory tract, or other organs and is generally unassociated with any underlying disease.

Antibody Response to Bacteriophage ΦX 174 in Non-Mammalian Vertebrates.

Discussion and Summary After a single injection of 108–10 bacteriophage ΦX 174, the chicken, frog and goldfish were shown to produce approximately the same levels of neutralizing, rapidly sedimenting, γ-globulin antibodies as those previously obtained in analogously immunized mammals(5,6). Repeated injections of bacteriophage in the frog and goldfish, at intervals of 2–4 weeks, did not elicit an anamnestic antibody response. However, higher levels of antibody, mainly in the slowly sedimenting γ-globulin fraction were produced after immunization with bacteriophage in complete Freunds adjuvant, and, in the case of the goldfish, after further elevation of the environmental temperature to 32°C. Thus, in 3 classes of non-mammalian vertebrates a change was observed in the sedimentation properties of antibody γ-globulins produced during immunization. This change appeared similar to the replacement of 19S by 7S antibodies in the circulation of immunized mammals. These findings suggest that the mechanisms responsible for this phenomenon were present in the most recent common ancestors of terrestrial vertebrates and bony fish and that formation of rapidly sedimenting antibody is an integral and important part of the immune mechanism.