Experimental Observations of Laser-Driven Tin Ejecta Microjet Interactions.

The study of high-velocity particle-laden flow interactions is of importance for the understanding of a wide range of natural phenomena, ranging from planetary formation to cloud interactions. Experimental observations of particle dynamics are sparse given the difficulty of generating high-velocity flows of many particles. Ejecta microjets are micron-scale jets formed by strong shocks interacting with imprinted surfaces to generate particle plumes traveling at several kilometers per second. As such, the interaction of two ejecta microjets provides a novel experimental methodology to study interacting particle streams. In this Letter, we report the first time sequences of x-ray radiography images of two interacting tin ejecta microjets taken on a platform designed for the OMEGA Extended Performance (OMEGA EP) laser. We observe that the microjets pass through each other unattenuated for the case of 11.7±3.2 GPa shock pressures and jet velocities of 2.2±0.5 km/s but show strong interaction dynamics for 116.0±6.1 GPa shock pressures and jet velocities of 6.5±0.5 km/s. We find that radiation-hydrodynamic simulations of the experiments are able to capture many aspects of the collisional behavior, such as the attenuation of jet velocity in the direction of propagation, but are unable to match the full spread of the strongly interacting cloud.

Radiographic and histologic evaluation of platelet-rich plasma and bovine-derived xenograft combination in bilateral sinus augmentation procedure.

There is currently a great interest regarding the use of platelet-rich plasma (PRP) in combination with various bone graft materials in sinus lift procedures. The purpose of this study was to assess and compare the radiographic and histological results of sinus augmentation procedures following treatment with PRP/bovine-derived xenograft (BDX) vs. BDX/collagen membrane. Using a split mouth design, 10 patients, with ≤5 mm of residual alveolar bone in the vertical direction, were treated with PRP/BDX or BDX/collagen membrane. At 8 months after surgery, both graft materials led to a satisfactory increase in vertical dimensions of bone. Bone biopsies were taken from the augmented sites during the implant placement. Histological analysis demonstrated that majority of the trabecula contained orderly layered lamellar bone in the PRP/BDX group, whereas mainly woven bone with a haphazard arrangement of collagen fibers were noticed in the BDX /collagen membrane group. It can be concluded that both combinations resulted with a satisfactory bone height, but more prominent and mature bone formation was observed at sites treated with PRP/BDX.

[Root resection in the era of dental implants].

The treatment of furcation defects is one of the most challenging aspects of periodontal therapy. The periodontal therapist can utilize various treatment modalities, including: non surgical root debridement, local drug delivery, open flap surgery, tunneling, root resection, guided tissue regeneration and extraction. Each treatment method has its advantages and disadvantages. The dilemmas concerning tooth prognosis and prosthetic considerations in such cases are especially demanding, especially in comparison to implant therapy. The clinician's decision in these situations must comply with the objective condition in the particular case based on his abilities and knowledge and the patient expectations. In order to evaluate the relevancy of root resection procedures we reviewed the literature focusing on root resection therapy prognosis as well as the different therapeutic alternatives for furcated molar teeth. Root resection treatment guidelines, indications and contraindication are presented along with clinical examples. Root resection is currently a relevant treatment modality for furcation defects. By using proper case selection, good surgical technique, proper prosthetic treatment and good periodontal supportive care, a good 5 year prognosis can be achieved and complications can be minimized. In light of the growing literature concerning dental implants complications, extraction and dental implant placement should be recommended as the last option when all other conservative options cannot be used, or following their failure.

Immunocytochemical localization of the lens main intrinsic polypeptide (MIP26) in communicating junctions.

Plasma membranes of vertebrate lens fiber cells contain a major intrinsic polypeptide with an apparent molecular weight of 26,000 (MIP26). These plasma membranes are extremely rich in communicating junctions, and it has been suggested that MIP26 is a component of them. MIP26 was purified from cow lenses using preparative SDS gel electrophoresis followed by hydroxylapatite column chromatography. From gel electrophoresis patterns and aggregational properties it was concluded that the MIP26 preparation was homogeneous. The purified MIP26 was used to produce monospecific antibodies in rabbits as assessed by double immunodiffusion and crossed immunoelectrophoresis of purified MIP26 and solubilized lens plasma membranes against the antiserum. Indirect immunocytochemical studies were performed on open and closed lens plasma membrane vesicles by incubation in anti-MIP antiserum followed by ferritin-conjugated goat antirabbit IgG. The conjugate bound unequivocally to lens communicating junctions, indicating that MIP26 is a component of these structures.

Immunocytochemical localization of the main intrinsic polypeptide (MIP) in ultrathin frozen sections of rat lens.

The in situ distribution of the 26-kdalton Main Intrinsic Polypeptide (MIP or MP 26), a putative gap junction protein in ocular lens fibers, was defined at the electron microscope level using indirect immunoferritin labeling of ultrathin frozen sections of rat lens. MIP was found distributed throughout the plasma membrane of the lens fiber cell, with no apparent distinction between junctional and nonjunctional membrane. MIP was not detectable in the basal or lateral plasma membrane of the lens epithelial cell, including the interepithelial cell gap junctions; nor was MIP detectable in the plasma membrane or gap junctions of the hepatocyte. Previous reports have indicated that the protein composition of the lens fiber cell junction differs from that of the hepatocyte gap junction. The evidence presented here suggests that the composition of the fiber cell junction and plasma membrane is also immunocytochemically distinct from that of its progenitor, the lens epithelial cell.

Tau-crystallin/alpha-enolase: one gene encodes both an enzyme and a lens structural protein.

tau-Crystallin has been a major component of the cellular lenses of species throughout vertebrate evolution, from lamprey to birds. Immunofluorescence analysis of the embryonic turtle lens, using antiserum to lamprey tau-crystallin showed that the protein is expressed throughout embryogenesis and is present at high concentrations in all parts of the lens. Partial peptide sequence for the isolated turtle protein and deduced sequences for several lamprey peptides all revealed a close similarity to the glycolytic enzyme enolase (E.C. 4.2.1.11). A full-sized cDNA for putative duck tau-crystallin was obtained and sequenced, confirming the close relationship with alpha-enolase. Southern blot analysis showed that the duck genome contains a single alpha-enolase gene, while Northern blot analysis showed that the message for tau-crystallin/alpha-enolase is present in embryonic duck lens at 25 times the abundance found in liver. tau-Crystallin possesses enolase activity, but the activity is greatly reduced, probably because of age-related posttranslational modification. It thus appears that a highly conserved, important glycolytic enzyme has been used as a structural component of lens since the start of vertebrate evolution. Apparently the enzyme has not been recruited for its catalytic activity but for some distinct structural property. tau-Crystallin/alpha-enolase is an example of a multifunctional protein playing two very different roles in evolution but encoded by a single gene.

Dietary restriction retards age-related loss of gamma crystallins in the mouse lens.

The soluble crystallins in lenses from diet-restricted and control mice of diverse ages (2, 11, or 30 months) were studied by high-performance liquid chromatography and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Results obtained with both methods suggest that dietary restriction decelerates age-related loss of soluble gamma crystallins.

A-ring substituted estrogens as inhibitors of the MXT transplantable mammary ductal carcinoma.

A-ring substituted estrogens have been examined as growth inhibitors of the hormone dependent MXT murine mammary tumor. Certain of these estrogen analogues inhibited the growth of newly implanted as well as established MXT tumors when administered either by s.c. or i.p. injections or by intubation. These compounds were nontoxic over a broad range of active levels. Amino and nitro groups, introduced at position-4 of estrone 3-methyl ether were particularly carcinostatic, a property not shared by 4-bromoestrone 3-methyl ether. In addition tumor inhibition was greatly diminished by placing the nitro group at the other ortho position (i.e., carbon-2). Evidence indicates that the A-ring substituted estrogens may function as growth inhibitors via the estrogen receptor mechanism in the case of 4-nitro- and 4-aminoestrone. The 3-methyl ethers of these compounds also blocked tumor growth, possibly through in vivo dealkylation leading to the free phenolic A-ring substituted estrogens. On the other hand, A-ring substituted 3-deoxyestrogens (particularly 4-nitro- and 4-aminoestratrien-17 beta-ol), which do not bind to receptor, were also excellent inhibitors of hormone dependent MXT breast tumors and therefore must express their activity by mechanisms other than that mediated by receptor. The A-ring substituted estrogens are unlike tamoxifen and diethylstilbestrol which (a) display toxicity at optimum inhibitory doses and (b) are inactive or marginally active in rodent breast cancer models.

Pharmacokinetics of 5-fluorouracil administered orally, by rapid intravenous and by slow infusion.

Pharmacokinetic studies of 5-fluorouracil (5-FUra) were performed on 18 patients divided into three groups: seven patients were given 5-FUra i.v. by rapid injection; five patients received the drug p.o.; and six patients were treated by continuous i.v. infusion for 96 hr. The results showed rapid i.v. injection to be manifested by high early levels of drug achieved both in plasma and bone marrow with a rapid fall afterwards. Administration of 5-FUra p.o. gave rise to erratic plasma values due to greater variability in absorption, whereas 96-hr i.v. infusions showed constant levels of the drug in plasma and significantly (50- to 1000-fold) less 5-FUra in bone marrow. The main difference observed between rapid injection and slow infusion in the kinetics of the drug was the very high level of 5-FUra reached by rapid injection in plasma and bone marrow, which was of short duration (min) when compared to the low sustained levels observed during infusion. This route-dependent pharmacokinetic profile is consistent with the reported absence of myelosuppression in prolonged infusion and may be related to the resultant lower levels of 5-FUra achieved in bone marrow.

In vitro biological evaluation of the R and S isomers of 1-(Tetrahydrofuran-2-yl)-5-fluorouracil.

The S isomer of Ftorafur was synthesized and the ability of the latter to inhibit growth of cultured human fibroblasts was determined relative to both the R isomer and the racemic mixture (Ftorafur) that is presently used clinically. No significant difference in the cytotoxic effects or the relative abilities to prevent an increase in cell numbers was observed with the three forms. Inhibition of DNA synthesis in murine L1210 leukemia cells by either isomer was observed only after prolonged (18-hr) exposure. The data suggest that Ftorafur is a repository form of 5-fluorouracil and that activity is manifested equally by both isomers.

Effects of 4-nitroestrone 3-methyl ether on dimethylbenz(a)anthracene-induced mammary tumors.

4-Nitroestrone 3-methyl ether has been shown to be an effective growth inhibitor of certain dimethylbenz(a)anthracene-induced rat mammary tumors in intact or ovariectomized rats. When administered at optimum levels (24 mg/kg daily), this A-ring-substituted estrone displayed no toxicity, slight estrogenicity, and an antitumor activity which was comparable to that of tamoxifen and nafoxidine and was surpassed only by ovariectomy or pharmacological doses of 17 beta-estradiol 3-benzoate. In addition, the appearance of mammary tumors was prevented when this estrogen derivative was administered to rats just prior to or after dimethylbenz(a)anthracene intubation. Unique to the action of the methyl ether of 4-nitroestrone on mammary tumors was the destruction of adenocarcinomas while permitting the appearance of fibroadenomas. Systemically, 4-nitroestrone 3-methyl ether brought about focal atrophy within the pituitary and ovaries while causing moderate hypertrophy of the uterus. Plasma prolactin was unaffected.

[Using acellular dermal matrix (ADM) allograft in periodontal surgery--a literature review and case reports].

The acellular dermal matrix (ADM) allograft is widely used in periodontal surgery. A MEDLINE search was performed for retrieving related articles. This article reviews ADM processing and mode of use, indications and contraindications, surgical techniques and clinical results for the treatment of localized and generalized gingival recessions with ADM and comparisons with other methods of treatment for root recessions. Two case reports are presented, illustrating the use of ADM and connective tissue grafts for root coverage.

Characterization and enzyme activity of argininosuccinate lyase/delta-crystallin of the embryonic duck lens.

Argininosuccinate lyase (ASL)/delta-crystallin, a major soluble protein of the transparent eye lens of birds and reptiles, is a mixture of tetramers comprising all possible combinations of two similar polypeptides (delta 1 and delta 2). Only the delta 2 polypeptide has ASL activity. In the present investigation we have purified each of the 5 major isoforms (delta A to delta E, pI 5.2 to 5.8) of delta-crystallin tetramers from the embryonic duck lens by isoelectric focussing and established by peptide sequencing that the delta 1 and delta 2 polypeptides are encoded in the previously identified, linked delta 1 and delta 2 genes, respectively. The relative amounts of the different tetramers in the 14-day-old embryonic lens were consistent with equal expression of the 2 delta-crystallin genes and no preference for assembly of the 2 delta polypeptides. The relative amount of ASL activity of the tetramers was a linear function of the relative amount of their delta 2 polypeptides, with delta A (only delta 1) lacking enzymatic activity altogether. delta B (3 delta 1:1 delta 2), delta C (2 delta 1:2 delta 2), delta D (1 delta 1:3 delta 2) and delta E (4 delta 2) all gave normal Michaelis-Menten kinetics for fumarate production from argininosuccinate at 40 degrees C and had a similar Km (average Km for mixture was 0.15 mM). delta E had a Km of 0.187 mM and a Vmax of 9 mumol/min per mg protein. Unlike bovine and like human ASL, both reported previously, embryonic duck ASL/delta-crystallin showed no evidence of cooperativity or activation by GTP. Each isoform had a similar far ultraviolet circular dichroism spectrum and thermal stability between 20 degrees C and 60 degrees C, with denaturation occurring at 65 degrees C. Our data suggest that gene duplication, structural modifications leading to greater thermal stability of the delta 1 and delta 2 polypeptides, and selective loss of ASL activity in the delta 1 polypeptide all occurred during the recruitment of ASL for a refractive role in the duck lens, resulting in the generation of ASL isoenzymes.

Cloning, sequencing and differential expression of alphaB-crystallin in the zebrafish, Danio rerio.

Here we report the cloning and expression of alphaB-crystallin from the zebrafish. 5'- and 3'-RACE was used to isolate a 900-bp transcript that contained insertions and deletions that differentiate it from both alphaA-crystallin and HSP-27. The deduced amino acid sequence of zebrafish alphaB-crystallin revealed that it lacked four residues in the C-terminus implicated in protein-protein interactions in other vertebrate species. In addition, the sequence contained two substitutions at sites implicated in phosphorylation in other vertebrate species. Northern analysis and semi-quantitative RT-PCR indicate that zebrafish alphaB-crystallin is expressed at extremely low levels outside of the lens.

The interaction of Cibacron Blue F3GA with troponin and its subunits.

Binding of troponin to Cibacron Blue F3GA-agarose column and its selective release from the gel in the presence of 0.5 M KCl provides the basis for a new purification method. The two-step procedure consists of isoelectric precipitation of tropomyosin and chromatography of the resultant crude troponin supernatant on Affi-Gel Blue column. Adsorption of troponin to the immobilized dye appears to occur through the troponin-T subunit. Troponin-I and troponin-C do not bind to the blue agarose column, whereas troponin-T binds to it very tightly. Binding of the dye to troponin-T prevents formation of troponin-T-troponin-C complex, but does not interfere with direct interaction of troponin-T with troponin-I. The activity of troponin in conferring calcium sensitivity on actomyosin ATPase is not affected by Cibacron Blue. Circular dichroism and difference absorption measurements of complexes of the blue dye with troponin and its subunits reveal the presence of a tight binding site on whole troponin and on troponin-T (KA greater than or equal to 10(6) M). The existence of weak binding sites for the dye on troponin and all of its subunits is deduced from difference absorption studies. Cibacron Blue appears to be a sensitive probe for subunit interactions in troponin.

Partial dissociation and renaturation of embryonic chick delta-crystallin. Characterization by ultracentrifugation and circular dichroism.

1. delta-Crystallin from 15-day-old embryonic chick lenses was characterized by circular dichrosim (CD) spectroscopy. Examination by CD spectroscopy in the far ultraviolet (190-250 nm) demonstrated that the secondary structure of delta-crystallin has at least 75% alpha-helix; the delta-crystallin subunits dissociated in sodium dodecyl sulfate retain their alpha-helical content. This appreciable alpha-helical content of delta-crystallin contrasts with the absence of alpha-helix in other lens crystallins. 2. As judged by CD spectroscopy in the near ultraviolet (250-320 nm) the tertiary structure of embryonic delta-crystallin is not readily disrupted by environmental changes, such as NaCl, KSCN or the non-ionic detergent Emulphogene BC 720, and is stable to temperature fluctuation between 2 and 56 degrees C. 3. Experiments were directed towards deaggregation and renaturation of the four subunits of embryonic delta-crystallin by treatment with urea or guanidine hydrochloride. The native tertiary structure of delta-crystallin was lost above 4 M urea or 2 M guanidine hydrochloride, as judged by CD spectroscopy in the near ultraviolet. Ultracentrifugation at sedimentation equilibrium showed that in 4 M urea delta-crystallin dissociates into dimeric subunits, while in 2 M guanidine hydrochloride delta-crystallin exists as a mixture of dimeric and tetrameric subunits. Dialysis of delta-crystallin from 4 M urea resulted in reaggregation of the subunits into tetramers, about 50% of which showed native tertiary structure. Dialysis from 2 M guanidine hydrochloride also resulted in tetramer formation, and about 35% was recovered with native conformation. Removal of denaturant by dialysis produced no native teritary structure after treatment with 8 M urea, but about 15% native conformation after treatment with 6 M guanidine hydrochloride.

Evidence that alpha-crystallin prevents non-specific protein aggregation in the intact eye lens.

The ocular lens is a transparent organ comprised of a highly concentrated and highly ordered matrix of structural proteins, called crystallins, which are probably the longest lived proteins of the body. Lens transparency is dependent upon maintenance of the short range order of the crystallin matrix. This transparency must be maintained for decades in the absence of normal protein synthesis or repair capacity. We present evidence here that alpha-crystallin, one of the major lens proteins, plays a central role in vivo in stabilizing the other crystallins and preventing uncontrolled aggregation of these progressively modified and aging molecules. alpha-Crystallin has previously been shown to suppress non-specific aggregation of denaturing proteins in simple binary systems through a chaperone-like activity. Our studies using soluble homogenates of monkey lenses demonstrate a strong resistance to heat induced non-specific aggregation when the complete complement of crystallins is present; in contrast, if alpha-crystallin is selectively removed prior to heating, the remaining crystallins undergo extensive non-specific aggregation as indicated by light scattering. When alpha-crystallin is present it complexes with denaturing proteins forming a soluble heavy molecular weight (HMW) fraction but no insolubilization is observed, while when alpha-crystallin is absent there is heavy insolubilization and no HMW formed. When intact monkey lenses were heated it could be demonstrated that soluble HMW was generated. Similar HMW protein appears in vivo in the human lens as a function of age. These findings suggest that the soluble HMW protein present in the human lens is the product of the chaperone-like function of alpha-crystallin and that under physiological conditions alpha-crystallin inhibits the uncontrolled aggregation of damaged proteins, thereby preventing the formation of light scattering centers and opacification of the lens.

Major intrinsic polypeptide of lens membrane. Biochemical and immunological characterization of the major cyanogen bromide fragment.

A protein of Mr 26000 has been shown to be the major component of eye-lens junctions, which are similar but not identical to the gap junctions of liver and other tissues. Cyanogen bromide cleavage of the Mr 26000 polypeptide from bovine lenses yields a major fragment of Mr 15000 (fragment 1). However, if the junctions are first treated with trypsin or carboxypeptidase Y, cyanogen bromide treatment yields a fragment of reduced molecular weight. Since protease treatment has been shown to cleave residues almost exclusively from the carboxy-terminal end of the Mr 26000 polypeptide, it follows that fragment 1 represents the carboxy-terminal half of this molecule, part of which is exposed to proteolytic attack outside the membrane. This latter result is corroborated by the fact that antisera which recognize both the Mr 26000 polypeptide and fragment 1 fail to do so after preadsorption with intact membranes. In addition, comparative amino acid and partial sequence analyses of the Mr 26000 polypeptide and fragment 1 indicate that fragment 1 is more hydrophilic in character, suggesting that much of the amino-terminal half of the Mr 26000 polypeptide is buried within the lipid bilayer.

Presence of a phospholipase D (PLD) distinct from PLD1 or PLD2 in human neutrophils: immunobiochemical characterization and initial purification.

Utilizing the transphosphatidylation reaction catalyzed by phospholipase D (PLD) in the presence of a primary alcohol and the short-chain phospholipid PC8, we have characterized the enzyme from human neutrophils. A pH optimum of 7.8-8.0 was determined. PIP(2), EDTA/EGTA, and ATP were found to enhance basal PLD activity in vitro. Inhibitory elements were: oleate, Triton X-100, n-octyl-beta-glucopyranoside, divalent cations, GTPgammaS and H(2)O(2). The apparent K(m) for the butanol substrate was 0.1 mM and the V(max) was 6.0 nmol mg(-1) h(-1). Immunochemical analysis by anti-pan PLD antibodies revealed a neutrophil PLD of approximately 90 kDa and other bands recognized minimally by anti-PLD1 or anti-PLD2 antibodies. The 90-kDa protein is tyrosine-phosphorylated upon cell stimulation with GM-CSF and formyl-Met-Leu-Phe. Protein partial purification using column liquid chromatography was performed after cell subfractionation. Based on the enzyme's regulatory and inhibitory factors, and its molecular weight, these data indicate an enzyme isoform that might be different from the mammalian PLD1/2 forms described earlier. The present results lay the foundation for further purification of this granulocyte PLD isoform.

Studies on bovine adrenal estrogen sulfotransferase. III. Facile synthesis of 3'-phospho- and 2'-phosphoadenosine 5'-phosphosulfate.

A practical synthesis of 3'-phosphoadenosine 5'-phosphosulfate (IV) in yields of 68-72% from adenosine 2',3'-cyclic phosphate 5'-phosphate (II) is described. Reaction of II with triethylamine-N-sulfonic acid affords adenosine 2',3'-cyclic phosphate 5'-phosphosulfate (III) which, on treatment with ribonuclease-T2, provides IV. Spleen phosphodiesterase, on the other hand, converts III to 2'-phosphoadenosine 5'-phosphosulfate (V). The biological activity of IV, measured by sulfate transfer to [6,7-3H2]estrone as mediated by bovine adrenal estrone sulfotransferase (3'-phosphoadenylyl-sulfate:estrone 3-sulfotransferase, EC 2.8.2.4), is identical with that obtained with a sample of IV prepared by an established biochemical procedure. By contrast, V exhibits approximately one-third the activity of the natural isomer.