Salivary defense factors in herpes simplex virus infection

Saliva may contribute to a lowering of the infectious herpes simplex virus (HSV) dose during transmission and consequently abrogate infection or lead to decreased reactivation. To test this hypothesis, we assayed saliva for innate defense factors, immunoglobulin content, and the capacity to interfere with HSV infection. Serum or salivary anti-HSV IgG levels did not correlate with control of recurrent labial herpes (RLH) and were significantly higher in subjects with RLH compared with asymptomatic seropositive subjects. Although no differences in levels or output rate of innate defense factors between the groups were observed, the salivary neutralizing activity correlated with lactoferrin and hypothiocyanite concentrations in the asymptomatic seropositive group. Our results suggest that saliva contains factors, in addition to anti-HSV immunoglobulins, that neutralize HSV and may indirectly contribute to the control of RLH.     

Rapid neural growth: Calcitonin gene-related peptide and substance P-containing nerves attain exceptional growth rates in regenerating deer antler

Deer antler is a unique mineralized tissue which can produce very high growth rates of > 1 cm/day in large species. On completion of antler growth, the dermal tissues which cover the antler are shed and the underlying calcified tissue dies. After several months the old antler is discarded and growth of a new one begins. It is known that deer antlers are sensitive to touch and are innervated. The major aims of this study were to identify and localize by immunohistochemical techniques the type of innervation present, and to find out whether nerve fibres could exhibit growth rates comparable to those of antler. We have taken tissue sections from the tip and shaft of growing Red deer (Cervus elaphus) antlers at three stages of development; shortly after the initiation of regrowth, the rapid growth phase, androduct 9.5 (a neural cytoplasmic protein), neurofilament triplet proteins (a neural cytoskeletal protein), substance P and calcitonin gene-related peptide (both of which are present in and synthesized by sensory neurons) showed the presence of immunoreactive nerve fibres in dermal, deep connective and perichondrial/periosteal tissues at all stages of antler growth. The sparse distribution of vasoactive intestinal polypeptide-like immunoreactivity was found in dermal tissue only at the earliest stage of antler development. Nerve fibres immunoreactive to neuropeptide Y, C-flanking peptide of neuropeptide Y and tyrosine hydroxylase, all present in postganglionic sympathetic nerves, were not observed at any stage of antler growth. Nerves expressing immunoreactivity for any of the neural markers or peptides employed could not be found in cartilage, osteoid or bone.

These results show that antlers are innervated mainly by sensory nerves and that nerves can attain the exceptionally high growth rates found in regenerating antler.     

Distribution of nerve endings and sensory neuropeptides in rat synovium, meniscus and bone.

The nervous system collects information from the outer world by specific senses and from the interior milieu by somatic senses. This information is processed and stored in memory and affects various bodily functions through the efferent arm of the nervous system. The efferent chemical neuropeptide message is transported intra-axonally to the site of action, which imparts site-specificity to the peripheral, paracrine neuropeptide effects. In the present study, immunohistochemistry using the immunoperoxidase method with nickel amplification was applied to visualize the topographical distribution of articular nerve fibres and nerve endings using the markers PGP 9.5 and synaptophysin, respectively. Furthermore, to get a comprehensive idea of the sensory innervation of the articular and para-articular tissue, antisera to calcitonin gene-related peptide (CGRP) and substance P were employed. Samples were collected after fixation by perfusion followed by immersion in fixative and decalcification by a special method, which also allows studies of the bone innervation. PGP 9.5- and synaptophysin-immunoreactive type IVa and IVb nerve fibres and endings were found in the synovial lining and sublining tissue and in the vascularized peripheral parts of the menisci. Furthermore, periosteum, bone marrow and the epiphyseal growth plates were also innervated, whereas innervation of the diaphyseal and metaphyseal bone was more sparse. PGP 9.5- and synaptophysin-immunoreactive nerves were also characterized by their CRGP, and to some extent, substance P content. Because of their distribution, the peripheral peptide-containing type IVa and IVb nerve fibres and nerve endings are in a position to participate in the pathogenesis of arthritis, including aspects of nociception, tissue remodelling and neurogenic inflammation.     

Serum level of CNTF is elevated in patients with amyotrophic lateral sclerosis and correlates with site of disease onset

We measured serum levels of neurotrophic cytokines ciliary neurotrophic factor (CNTF) and leukaemia inhibiting factor (LIF) in 96 patients either with familial amyotrophic lateral sclerosis (FALS, n  = 18) or sporadic ALS (SALS, n  = 78) and in 27 inflammatory neurological controls (13 multiple sclerosis and 14 Guillain-Barré syndrome) and in 27 healthy controls. Serum level of CNTF was significantly higher in ALS patients than in inflammatory neurological controls or healthy controls, and significantly higher in patients with ALS onset from upper or lower extremities than in patients with a purely bulbar onset of the disease. Serum CNTF levels did not significantly differ between patients with FALS and SALS, and it did not correlate with the age of onset or duration of the disease. No detectable serum levels of LIF were observed in the patient groups or in the healthy controls.     

Time-dependent sensory nerve ingrowth into a bone conduction chamber

We studied time-dependent ingrowth of sensory nerve fibers into a bone defect in a rat bone conduction chamber model. In 10 male Sprague Dawley rats, a titanium chamber was implanted bilaterally in the proximal tibiae, representing an experimental bone defect. To mimic a clinical situation, the chambers were filled with a fresh blood clot. After 1, 2, 4, 6 and 8 weeks, 2 rats were fixed in vivo at each time before removal of specimens, and histological and immunohistochemical analyses. We used antisera against protein gene product 9.5, neural growth-associated protein 43/B-50, calcitonin gene-related peptide, and substance P, to locate regenerating sensory nerve fibers in the chamber. During bone defect healing, hematoxylin/eosin sections showed that new bone grew in through the ingrowth openings in the chamber, gradually filling it and replacing the blood clot. At 1 and 2 weeks after implantation, no nerve fibers could be detected. At 4, 6 and 8 weeks, however, small numbers of nerve fibers were seen in 8 of 11 specimens. The nerve fibers were located mainly in the dense fibrous tissue in close proximity to the new bone, and in some cases within the new forming bone. In this chamber model, the periosteum is not in contact with the bone ingrowth openings, and all ingrowing nerve fibers thus originated from the cortical bone, endosteum or bone marrow. We speculated that these late ingrowing sensory nerve fibers may actively participate in bone repair.     

Correlation Between Ciliary Beat Frequency and the Structure of Ciliated Epithelia in Pathologic Human Nasal Mucosa

The structure and function of ciliated epithelium were studied in 44 human nasal mucosa samples using a photoelectric method and scanning electron microscopy (SEM). The ciliary beat frequency (CBF) of cases with recurrent or chronic sinusitis was 9.1 ± 5.4 Hz. In eight of the samples (18.2%) no ciliary activity was detected. The amount of ciliated cells, the orientation of cilia, epithelial metaplasia, and secretion were found to be explanatory factors accounting for the decreased ciliary activity. Ciliary disorientation and a lack of ciliated cells in SEM correlated with low ciliary activity. In cases where sinusitis secretion was not seen, the CBF was slower than in cases with mucus or mucopurulent secretion. Sinusitis with disoriented cilia, a loss of ciliated cells, and a lack of mucosal secretion is associated with a decreased CBF. This may lead to impaired mucociliary clearance and increase the risk of recurrent and chronic sinusitis.     

Pseudosynovial fluid from loosened total hip prosthesis induces osteoclast formation

Interface tissue between the bone and loosening total hip implant is acidic and highly osteolytic. It is characterized by the formation of cathepsin K positive foreign body giant cells. Similar structures to those found in the normal joint surround the artificial hip joint. Cells in synovial membrane of the artificial hip generate synovial fluid that is called pseudosynovial fluid. Interface tissue fibroblasts are able to produce receptor activator of NF-kappaB ligand (RANKL), which can induce osteoclastogenesis during the loosening process. Western blot analysis indicated that RANKL is present in the pseudosynovial fluid. Pseudosynovial fluid induced cultured peripheral blood mononuclear cells to form multinuclear TRAP positive giant cells. In the presence of osteoprotegerin, the soluble RANKL decoy receptor, the number of TRAP positive multinuclear cells was reduced to half (p < 0.05). The multinuclear cells induced with pseudosynovial fluid contained active cathepsin K protein and were capable of bone matrix resorption in vitro. The cells were shown to express osteoclast phenotype markers, such as mRNA for cathepsin K, TRAP, and calcitonin receptor. It is therefore apparent that pseudosynovial fluid from patients with aseptic loosening of total hip prosthesis contains a potent osteoclastogenic factor RANKL that further suggests a favorable environment for osteoclast formation in the peri-implant tissues. It is thus concluded that suppression of RANKL activity may be beneficial in terms of increasing the lifetime of total hip prostheses.     

Effects of two multi-step self-etch primer/adhesives on apoptosis in human gingival fibroblasts in vitro

Various in vitro studies have shown induction of apoptosis by monomers incorporated to dental restorative materials and adhesive resins, while information regarding the effect of monomer combinations as commercially available products on apoptosis is limited. The aim of this study was to investigate the effects of two multi-step self-etch primer/adhesive systems on apoptosis of cultured primary human gingival fibroblasts. Cells were treated up to 48 h with Clearfil SE Bond (Kuraray, Japan) and FL Bond (Shofu, Japan) at 1:1000 v:v ratio to determine cell proliferation, using 0.02 mM staurosporine as positive control. Apoptosis was assessed using propidium iodide/acridine orange (PI/AO) staining, compared to nontreated controls. When compared to FL Bond, exposure of gingival fibroblasts to Clearfil SE Primer and Clearfil SE Bond resulted in a higher degree of cell proliferation. PI/AO staining revealed typical morphological features of apoptosis in FL Bond and Staurosporine groups, while some cells cultured in the presence of primer and adhesive components of Clearfil SE Bond showed nuclear fragmentation, indicative of early apoptosis. Our results indicate that apoptotic potential of the multi-step self-etch adhesives were material-dependent within the 48 h test period.     

Mechanical strain stimulates nitric oxide production by rapid activation of endothelial nitric oxide synthase in osteocytes.

Previous studies have indicated that physiological levels of dynamic mechanical strain produce rapid increases in nitric oxide (NO) release from rat ulna explants and primary cultures of osteoblast-like cells and embryonic chick osteocytes derived from long bones. To establish the mechanism by which loading-induced NO production may be regulated, we have examined: nitric oxide synthase (NOS) isoform mRNA and protein expression, the effect of mechanical loading in vivo on NOS mRNA expression, and the effect of mechanical strain on NO production by bone cells in culture. Using Northern blot analyses, in situ hybridization, and immunocytochemistry we have established that the predominant NOS isoform expressed in rat long bone periosteal osteoblasts and in a distinct population of cortical bone osteocytes is the endothelial form of NOS (eNOS), with little or no expression of the inducible NOS or neuronal NOS isoforms. In contrast, in non-load-bearing calvariae there are no detectable levels of eNOS in osteocytes and little in osteoblasts. Consistent with these observations, ulnar explants release NO rapidly in response to loading in vitro, presumably through the activation of eNOS, whereas calvarial explants do not. The relative contribution of different bone cells to these rapid increases in strain-induced NO release was established by assessment of medium nitrite (stable NO metabolite) concentration, which showed that purified populations of osteocytes produce significantly greater quantities of NO per cell in response to mechanical strain than osteoblast-like cells derived from the same bones. Using Northern blot hybridization, we have also shown that neither a single nor five consecutive daily periods of in vivo mechanical loading produced any significant effect on different NOS isoform mRNA expression in rat ulnae. In conclusion, our results indicate that eNOS is the prevailing isoform expressed by cells of the osteoblast/osteocyte lineage and that strain produces increases in the activity of eNOS without apparently altering the levels of eNOS mRNA.     

Herpes simplex virus DNA in amniotic fluid without neonatal infection.

Twenty-one pregnant women were studied to determine the distribution of herpes simplex virus (HSV): 10 had symptomatic genital herpes, including 1 with primary cervical HSV infection, and 11 had asymptomatic genital herpes. Samples from vesicles, the cervix, and amniotic fluid (AF) were analyzed with 2 separate polymerase chain reaction (PCR) tests and with viral culture. For newborns, clinical examinations and pharyngeal HSV cultures were performed, and cord blood IgM antibodies to HSV were measured. HSV DNA was present in the AF of 3 women with symptomatic HSV infection, but all cultures were negative. HSV was detected more often with PCR than with culture, in both vesicle and cervical samples. For the asymptomatic group, all AF samples were negative, whereas 4 cervical samples were positive by PCR (none were positive by culture). All isolates were HSV type 2. All infants were healthy, and none had cord blood IgM antibodies to HSV, including those with PCR-positive AF.