ABSTRACT
Shortening the dry period improves postpartum energy balance, which has potential positive effects on metabolic health. This concept has been mainly studied in Holstein cows. The aim of this study was to assess the effect of a short dry period of 4 wk, compared with a standard dry period of 8 wk, on the metabolic status, progesterone profiles, health, and colostrum quality of dairy cows of 2 breeds, Swedish Red (SR) and Swedish Holstein (SH), not treated with antibiotics at dry off. The IgG uptake in calves was also studied to reflect the colostrum quality when shortening the dry period. Cows of both SH and SR were blocked by breed and parity and then randomly allocated to a short dry period of 4 wk (4W, n = 43) or a conventional dry period of 8 wk (8W, n = 34). Blood samples were collected wk -8, -4, -2, -1 and 1, 2, 3, 4, 6, 9, and 12 relative to calving. Prepartum, cows with a 4-wk dry period had higher concentrations of nonesterified fatty acids and lower concentrations of insulin-like growth factor-1 and insulin than 8W cows. Postpartum, plasma concentration of nonesterified fatty acids was lower, whereas plasma insulin and insulin-like growth factor-1 tended to be higher for 4W cows than for 8W cows. Plasma concentration of ß-hydroxybutyrate did not differ between dry period lengths. Swedish Holstein cows with a 4W dry period responded with a lower concentration of insulin prepartum than SR and SH on an 8W dry period. The dry period length had no effect on the proportion of disturbed progesterone profiles; disturbed progesterone profiles occurred in 30% of the 4W cows and 47% of the 8W cows. In this trial, only 48.8% of the SR cows had a normal progesterone profile, which differed from the SH where 76.5% had a normal profile. Fertility-related diseases (endometritis, pyometra, anestrus, ovarian cyst) did not differ between the 2 dry period groups: 21% in the 8W group versus 12% in the 4W group, whereas mastitis tended to be more common: 26% of the 4W cows versus 9% of the 8W cows. A short dry period resulted in less colostrum but with a higher content of protein and somatic cell count. Calves were fed colostrum from their dam, and the IgG and total protein in plasma did not differ between calves to mothers with different a dry period length. Shortening the dry period could improve metabolic status in cows of both SH and SR breed postpartum, without compromising the colostrum quality. Health and progesterone profiles were not affected by the dry period length for SH or SR in this study.
Subject(s)
Cattle/physiology , Colostrum/metabolism , Fertility , Lactation/physiology , Mammary Glands, Animal/physiology , 3-Hydroxybutyric Acid/blood , Animals , Breeding , Diet/veterinary , Energy Metabolism , Fatty Acids, Nonesterified/blood , Female , Insulin/blood , Insulin-Like Growth Factor I/analysis , Milk/chemistry , Milk/metabolism , Parity , Postpartum Period , Pregnancy , Progesterone/analysis , Species Specificity , SwedenABSTRACT
The present study investigated whether delaying the first feeding of colostrum affected ileum and colon mucosa-associated microbiota in calves. Twenty-seven male Holstein calves were randomly assigned to 1 of 3 groups, fed colostrum at 45 min, 6 h, and 12 h after birth, respectively. Ileum and colon mucosa were collected at 51 h after birth, and their associated microbial profiles were assessed using amplicon sequencing. Both ileum and colon mucosa-associated microbiota were predominated by genus Escherichia-Shigella. The negative correlation between the molar proportion of short-chain fatty acids (SCFA) and ileum mucosa-associated opportunistic pathogens, and the positive correlation between the molar proportion of SCFA and colon mucosa-associated beneficial bacteria, suggest that SCFA might play an important role in maintaining the gut health of 2-d-old calves. A higher relative abundance of ileum mucosa-associated Enterococcus and Streptococcus was detected when the first colostrum feeding was delayed for 12 h. The relative abundance of colon mucosa-associated Lactobacillus tended to be lower in calves fed colostrum 12 h than those under the other 2 treatments, whereas that of Faecalibacterium tended to be lower in calves fed colostrum immediately after birth than those fed colostrum 6 and 12 h after birth, respectively. Our findings suggest that delayed first colostrum feeding affects the establishment of ileum and colon mucosa-associated bacteria, which may have long-term effects on gut health of calves.
Subject(s)
Animals, Newborn/microbiology , Cattle/microbiology , Colostrum/metabolism , Fatty Acids, Volatile/analysis , Gastrointestinal Microbiome , Animals , Animals, Newborn/physiology , Cattle/physiology , Colon/microbiology , Enterococcus/classification , Enterococcus/growth & development , Escherichia/classification , Escherichia/growth & development , Female , Ileum/microbiology , Intestinal Mucosa/microbiology , Male , Random Allocation , Shigella/classification , Shigella/growth & development , Streptococcus/classification , Streptococcus/growth & development , Time FactorsABSTRACT
This pilot study introduces a novel vibratory stimulation-based occlusal splint (VibOS) for management of pain related to temporomandibular disorders (TMD). The study sample consisted of 10 patients (mean age: 40·5 ± 13·7 years, male/female: 3/7) who were using stabilisation splints for more than 2 months prior to the study onset and still complained of pain. Patients utilised the active and inactive VibOS during 15 days in a crossover designed clinical trial. The analysed variables were self-reported VAS pain levels and number of painful sites to palpation (PSP). Statistical analysis was performed with repeated measures anova. At baseline, mean VAS pain levels for group I and II were 45·6 ± 21·0 mm and 37·4 ± 16·3 mm, respectively. Comparison between these baseline values showed no statistical difference (P > 0·05, unpaired t-test). In group I, the inactive VibOS caused a slight increase in VAS pain levels, whereas the active VibOS promoted a significant decrease in VAS pain levels and PSP (P < 0·01). In group II, which received the active VibOS first, a significant decrease in VAS levels (P < 0·05) and in PSP (P < 0·01) was observed. No significant decrease in VAS pain levels or PSP (P > 0·05) was observed with the use of the inactive VibOS. In conclusion, this study demonstrated a good tendency of this novel VibOS in the alleviation of painful symptoms related to TMD after a 15-day management period compared to control VibOS.
Subject(s)
Arthralgia/therapy , Facial Pain/therapy , Occlusal Splints , Temporomandibular Joint Dysfunction Syndrome/therapy , Vibration/therapeutic use , Adolescent , Adult , Arthralgia/etiology , Cross-Over Studies , Facial Pain/etiology , Female , Humans , Joint Dislocations/complications , Male , Middle Aged , Pain Measurement , Pilot Projects , Temporomandibular Joint Disc/injuries , Temporomandibular Joint Dysfunction Syndrome/complications , Treatment OutcomeABSTRACT
This retrospective study identified the risk factors for fracture of veneering materials and screw loosening of implant-supported fixed partial dentures in partially edentulous cases. The study group included a total of 182 patients who were installed 219 suprastructures at the Fixed Prosthodontic Clinic of Okayama University Dental Hospital between February 1990 and March 2005 and were subdivided in two subgroups: 120 patients (149 facing suprastructures) were included in the subgroup to investigate the risk factors of fracture of veneering materials, and 81 patients (92 suprastructures) were included in the subgroup to identify the risk factors of abutment screw loosening. Each patient was followed up from the day of suprastructure installation until March, 2005. A Cox proportional hazards regression model was used to identify the risk factors related to technical complications, and eight factors were regarded as candidate risk factors. Screw retention was the significant risk factor for fracture of veneering materials, whereas connection of suprastructures with natural tooth was the significant risk factor for screw loosening. It was suggested that screw retention was a significant risk factor for the fracture of veneering materials, and connection of suprastructures with natural tooth was a significant risk factor for screw loosening. Future studies, involving dynamic factors (e.g. bruxism) as predictors as well, are more helpful to discuss the risk factor of fracture of veneering materials and screw loosening.
Subject(s)
Bone Screws/adverse effects , Dental Prosthesis, Implant-Supported/adverse effects , Dental Restoration Failure/statistics & numerical data , Dental Veneers/adverse effects , Jaw, Edentulous, Partially/rehabilitation , Dental Abutments , Female , Follow-Up Studies , Humans , Incidence , Male , Middle Aged , Retrospective Studies , Risk Factors , Time Factors , Treatment OutcomeABSTRACT
Parafunctional habits, such as prolonged clenching and bruxism, have been associated with dysfunctional overloading in the temporomandibular joint (TMJ). In this study, stress distributions in the TMJ were analysed during prolonged clenching, using three-dimensional finite element (FE) models of the TMJ with and without disc displacement. The aim of this study was to investigate stress distribution of the cartilaginous tissues in the TMJ with and without disc displacement. Finite element models were developed on the basis of magnetic resonance images from two subjects with and without anterior disc displacement. Condylar movements recorded during a 5-min clenching were used as the loading condition for stress analysis. In the asymptomatic model, the highest von Mises stresses were located in the lateral area (4·91 MPa) of the disc surfaces, and after 5-min clenching, the higher stresses were still located at the lateral area (3·65 MPa). In all the cartilaginous tissues, 30-50% of stress reduction occurred during 5-min clenching. In contrast, the von Mises stress in the cartilaginous tissues of the symptomatic model with disc displacement was markedly lower, compared with the asymptomatic model. However, in the condylar cartilage, stress relaxation during clenching was not recognised. Furthermore, relatively high stresses were observed in the retrodiscal tissues throughout clenching. The present results indicate that disc position could be involved in the stress distribution of the TMJ components during prolonged clenching.
Subject(s)
Finite Element Analysis , Joint Dislocations/physiopathology , Muscle Contraction/physiology , Temporomandibular Joint Disc/physiopathology , Temporomandibular Joint Disorders/physiopathology , Temporomandibular Joint/physiopathology , Adult , Algorithms , Biomechanical Phenomena , Cartilage, Articular/pathology , Cartilage, Articular/physiopathology , Computer Simulation , Elastic Modulus , Female , Friction , Humans , Imaging, Three-Dimensional/methods , Joint Dislocations/pathology , Magnetic Resonance Imaging , Mandibular Condyle/pathology , Mandibular Condyle/physiopathology , Masticatory Muscles/physiopathology , Models, Biological , Stress, Mechanical , Temporal Bone/pathology , Temporal Bone/physiopathology , Temporomandibular Joint/pathology , Temporomandibular Joint Disc/pathology , Temporomandibular Joint Disorders/pathology , Young AdultABSTRACT
Occlusal dysesthesia refers to a persistent complaint of uncomfortable bite sensation with no obvious occlusal discrepancy. This systematic review aimed to draw a picture of such patients, to present an agreement of previously reported diagnostic criteria and to analyse the evidence level of the recommended management approaches. An electronic search for all relevant reports on occlusal dysesthesia was thoroughly performed based on previous nomenclatures (e.g. phantom bite, occlusal hyperawareness) in PubMed and The Cochrane Library in July, 2011. A total of 84 reports were matched, among which only 11 studies were included after a two-step (abstract and detailed full-text revision) screening process. Additionally, a thorough manual review of reference lists of the included reports enabled the inclusion of two additional studies. Data analysis revealed that 37 occlusal dysesthesia patients presented a mean age of 51.7 ± 10.6 years and were predominantly women (male/female: 1/5.1) with symptom duration of more than 6 years (average: 6.3 ± 7.5 years) and with concomitant psychological disturbances (e.g. mood disorders, somatoform disorders, personality disorders). Only four authors presented diagnostic criteria for occlusal dysesthesia, which served as the basis for an agreement in the diagnostic criteria. Treatment approaches included psychotherapy, cognitive/behaviour therapy, splint therapy and prescription of anti-depressants or anti-anxiety drugs. Classification of evidence level of management approaches, however, revealed that most of them were expert opinions with single- or multiple-case report(s). Future studies are necessary for a deeper understanding of the mechanisms behind the occlusal dysesthesia symptoms, and consequently, for improvements in evidence-based management approaches.
Subject(s)
Malocclusion/psychology , Paresthesia/psychology , Somatoform Disorders/psychology , Adult , Female , Humans , Male , Malocclusion/etiology , Malocclusion/therapy , Middle Aged , Paresthesia/etiology , Paresthesia/therapy , Somatoform Disorders/complicationsABSTRACT
The purposes of this study were to determine whether a response shift was observable after partial denture treatment and to identify the predictors that influenced the response shift magnitude and direction. A total of 173 consecutive patients with no more than eight missing teeth who received implant-supported, fixed or removable partial dentures at Okayama University Dental Hospital were asked to complete a full-version Oral Health-Related Quality of Life (OHRQoL) questionnaire before (pre-test) and after treatment (post-test). Additionally, a short form (then-test) consisting of seven questions selected from the full version had its reliability verified and was utilised to retrospectively assess the pre-treatment OHRQoL status. The difference between the summary scores of the then-test and the pre-test determined the response shift magnitude and direction. The then-test mean score (22·9 ± 6·6) was significantly lower (worse OHRQoL) than that of the pre-test (26·4 ± 5·2). The response shift effect size was of moderate magnitude and negative direction (d = -0·78). A multiple regression analysis showed that age (younger patients) (P < 0·01), number of replaced teeth (fewer) (P < 0·01) and pre-test scores (lower) (P < 0·01) were the significant predictors for response shift. In conclusion, a response shift phenomenon with negative and moderate effect size was observed after partial denture treatment. The significant predictor variables were young age, fewer numbers of replaced teeth and lower pre-test scores.
Subject(s)
Denture, Partial/psychology , Jaw, Edentulous, Partially/rehabilitation , Oral Health , Patient Satisfaction , Quality of Life , Tooth Loss/rehabilitation , Adult , Aged , Aged, 80 and over , Female , Humans , Male , Middle Aged , Retrospective Studies , Surveys and Questionnaires , Treatment Outcome , Young AdultABSTRACT
Here we utilize chemical ecology as a tool to manipulate the biological system of a small, but highly venomous to humans, cubozoan jellyfish, Carukia barnesi. We trialled a range of chemical reagents including indole compounds, 9-cis-retinoic acid and lugols solution to induce metamorphosis between the polyp and medusa life stages. An optimum method was determined resulting in a 90% metamorphosis rate to healthy medusa by exposing the polyps to 1 µM of 5-methoxy-2-methylindole for 24 h. Of note is that chemical exposure time significantly impacts health and metamorphosis rates in this species. We also present a theoretical mechanism for the chemical/biological interactions occurring during metamorphosis. This is a significant methodological advancement which now enables rearing of this animal en mass in aquaria-a world first for this species-which will subsequently supply and facilitate venom research into this understudied jellyfish.
Subject(s)
Cnidaria , Cubozoa , Scyphozoa , Animals , Ecology , Metamorphosis, BiologicalABSTRACT
[This corrects the article DOI: 10.1016/j.toxcx.2021.100067.].
ABSTRACT
During orofacial tissue development, the anterior and posterior regions of the Meckel's cartilage undergo mineralization, while the middle region undergoes degeneration. Despite the interesting and particular phenomena, the mechanisms that regulate the different fates of Meckel's cartilage, including the effects of biomechanical cues, are still unclear. Therefore, the purpose of this study was to systematically investigate the course of Meckel's cartilage during embryonic development from a biomechanical perspective. Histomorphological and biomechanical (stiffness) changes in the Meckel's cartilage were analyzed from embryonic day 12 to postnatal day 0. The results revealed remarkable changes in the morphology and size of chondrocytes, as well as the occurrence of chondrocyte burst in the vicinity of the mineralization site, an often-seen phenomenon preceding endochondral ossification. To understand the effect of biomechanical cues on Meckel's cartilage fate, a mechanically tuned 3-dimensional hydrogel culture system was used. At the anterior region, a moderately soft environment (10-kPa hydrogel) promoted chondrocyte burst and ossification. On the contrary, at the middle region, a more rigid environment (40-kPa hydrogel) enhanced cartilage degradation by inducing a higher expression of MMP-1 and MMP-13. These results indicate that differences in the biomechanical properties of the surrounding environment are essential factors that distinctly guide the mineralization and degradation of Meckel's cartilage and would be valuable tools for modulating in vitro cartilage and bone tissue engineering.
Subject(s)
Cartilage , Mandible , Chondrocytes , Female , Humans , Osteogenesis , PregnancyABSTRACT
Venom research is often focussed on medical relevance, novel compounds and venom evolution, whilst studying the relationship between a venom and its environment - venom ecology - has been conducted to a lesser extent. Given the projected environmental changes envisioned to occur with global warming, it is pertinent now more than ever, to highlight this topic. Here we review literature examining the influence of ecological factors such as environmental temperature, salinity, ontogeny, geographic location and diet on cnidarian venoms. This review provides an exclusive focus on the cnidarian phylum and encompasses all available published, peer-reviewed literature to our knowledge regarding the ecological factors influencing venom. We find a startling lack of research into the effects of both environmental and biological factors on venoms, with very few to no studies available per category. Importantly, research does exist that suggest these ecological processes may influence other marine or terrestrial venoms, thus we recommend future research is needed to explore this concept in cnidarians.
ABSTRACT
Shortening the dry period (DP) has been proposed as a strategy to improve energy balance (EB) in cows in early lactation. This study evaluated the effects of shortening the DP on milk yield (MY), EB and residual feed intake (RFI) in two breeds; Swedish Red (SR) and Swedish Holstein (SH). Cows were blocked by breed and parity and then randomly assigned to one of two treatments; short DP of 4 weeks (4W, n=43) or conventional DP of 8 weeks (8W, n=34). Cows were kept and fed under the same conditions, except for the 4 weeks when the 4W group were still lactating prepartum and thus kept with the lactating cows. Milk yield and BW were recorded and body condition score (BCS) was rated from 10 weeks prepartum to 12 weeks postpartum. Dry matter intake (DMI) was recorded for lactating cows postpartum. Milk yield was reduced by 6.75 kg/day during the first 12 weeks postpartum (P<0.001) for the 4W cows compared with 8W cows, but there was no significant difference in total MY (3724 kg compared with 3684 kg, P=0.7) when the milk produced prepartum was included. Protein content was higher in 4W cows (3.42%) than in 8W cows (3.27%) (P<0.001) postpartum. In the 8W group, cows lost more BCS after calving (P<0.05). Cows of SR breed had higher BCS than cows of SH breed (SR=3.7, SH=3.2, P<0.001), but no differences in BW were found between breed and treatment. Energy balance was improved for cows in the 4W group (P<0.001), while feed efficiency, expressed as RFI, was reduced for 4W cows than for 8W cows (5.91 compared with -5.39, P<0.01). Shortening the DP resulted in improved EB postpartum with no difference between the breeds and no milk losses when including the milk produced prepartum.
Subject(s)
Cattle/physiology , Energy Metabolism , Milk/metabolism , Animals , Breeding , Diet/veterinary , Female , Lactation , Milk/chemistry , Parity , Postpartum Period , Pregnancy , Random AllocationABSTRACT
The Ets transcription factor PU.1 is a hematopoietic master regulator essential for the development of myeloid and B-cell lineages. As we previously reported, PU.1 sometimes represses transcription on forming a complex with mSin3A-histone deacetyl transferase-MeCP2. Here, we show an interaction between PU.1 and DNA methyltransferases, DNA methyltransferase (Dnmt)3a and Dnmt3b (Dnmt3s). Glutathione-S-transferase pulldown assay revealed that PU.1 directly interacted with the ATRX domain of Dnmt3s through the ETS domain. Dnmt3s repressed the transcriptional activity of PU.1 on a reporter construct with trimerized PU.1-binding sites. The repression was recovered by addition of 5-aza-deoxycitidine, a DNA methyltransferase inhibitor, but not trichostatin A, a histone deacetylase inhibitor. Bisulfite sequence analysis revealed that several CpG sites in the promoter region neighboring the PU.1-binding sites were methylated when Dnmt3s were coexpressed with PU.1. We also showed that the CpG sites in the p16(INK4A) promoter were methylated by overexpression of PU.1 in NIH3T3 cells, accompanied by a downregulation of p16(INK4A) gene expression. These results suggest that PU.1 may downregulate its target genes through an epigenetic modification such as DNA methylation.
Subject(s)
DNA (Cytosine-5-)-Methyltransferases/metabolism , DNA Methylation , Proto-Oncogene Proteins/pharmacology , Trans-Activators/pharmacology , Animals , Azacitidine/analogs & derivatives , Azacitidine/pharmacology , Chromatin Immunoprecipitation , Cyclin-Dependent Kinase Inhibitor p16/metabolism , DNA (Cytosine-5-)-Methyltransferases/genetics , DNA Methyltransferase 3A , Decitabine , Histone Deacetylase Inhibitors , Histone Deacetylases/metabolism , Hydroxamic Acids/pharmacology , Immunoprecipitation , Mice , NIH 3T3 Cells , Promoter Regions, Genetic , Proto-Oncogene Proteins/genetics , Trans-Activators/genetics , Transcription, Genetic , DNA Methyltransferase 3BABSTRACT
Rat 3Y1 cell lines that express either adenovirus type 12 E1A 13S mRNA or 12S mRNA in response to dexamethasone treatment were established by introduction of recombinant vector DNA containing the E1A 13S- or 12S-mRNA cDNA placed downstream of the hormone-inducible promoter of mouse mammary tumor virus. These cell lines were growth arrested, and the induction of cell cycle progression was analyzed by flow cytometry after switch on of the cDNA by the addition of dexamethasone. The results indicate that the 13S- or 12S-mRNA product alone has the ability to cause progression of the cell cycle at a similar rate. The simultaneous addition of epidermal growth factor accelerated the rate of cell cycle progression in the transition from the G0/G1 phase to the S phase.
Subject(s)
Cell Cycle , DNA/biosynthesis , Oncogene Proteins, Viral/physiology , Adenovirus Early Proteins , Animals , Cell Line , Cloning, Molecular , Dexamethasone/pharmacology , Epidermal Growth Factor/pharmacology , RNA, Messenger/genetics , RatsABSTRACT
Cell proliferation and differentiation are precisely coordinated during the development and maturation of the mammary gland, and this balance invariably is disrupted during carcinogenesis. Little is known about the cell-specific transcription factors that regulate these processes in the mammary gland. The mouse mammary epithelial cell line SCp2 grows well under standard culture conditions but arrests growth, forms alveolus-like structures, and expresses beta-casein, a differentiation marker, 4 to 5 days after exposure to basement membrane and lactogenic hormones (differentiation signals). We show that this differentiation entails a marked decline in the expression of Id-1, a helix-loop-helix (HLH) protein that inactivates basic HLH transcription factors in other cell types. SCp2 cells stably transfected with an Id-1 expression vector grew more rapidly than control cells under standard conditions, but in response to differentiation signals, they arrested growth and formed three-dimensional structures similar to those of control cells. Id-1-expressing cells did not, however, express beta-casein. Moreover, 8 to 10 days after receiving differentiation signals, they lost three-dimensional organization, invaded the basement membrane, and then resumed growth. SCp2 cells expressing an Id-1 antisense vector grew more slowly than controls; in response to differentiation signals, they remained stably growth arrested and fully differentiated, as did control cells. We suggest that Id-1 renders cells refractory to differentiation signals and receptive to growth signals by inactivating one or more basic HLH proteins that coordinate growth and differentiation in the mammary epithelium.
Subject(s)
Cell Differentiation/drug effects , DNA-Binding Proteins/pharmacology , Helix-Loop-Helix Motifs , Mammary Glands, Animal/drug effects , Repressor Proteins , Transcription Factors , Animals , Cell Line , DNA-Binding Proteins/metabolism , Epithelial Cells , Epithelium/drug effects , Inhibitor of Differentiation Protein 1 , Mice , Signal TransductionABSTRACT
p16CDKN2 specifically binds to and inhibits the cyclin-dependent kinases CDK4 and CDK6, which function as regulators of cell cycle progression in G1 by contributing to the phosphorylation of the retinoblastoma protein (pRB). Human cell lines lacking functional pRB contain high levels of p16 RNA and protein, suggesting a negative feedback loop by which pRB might regulate p16 expression in late G1. By a combination of nuclear run-on assays and promoter analyses in human fibroblasts expressing a temperature-sensitive simian virus 40 T antigen, we show that p16 transcription is affected by the status of pRB and define a region in the p16 promoter that is required for this response. However, the effect is not sufficient to account for the differences in p16 RNA levels between pRB-positive and -negative cells. Moreover, p16 RNA is extremely stable, and the levels do not change appreciably during the cell cycle. Primary human fibroblasts express very low levels of p16, but the RNA and protein accumulate in late-passage, senescent cells. The apparent overexpression of p16 in pRB-negative cell lines is therefore caused by at least two factors: loss of repression by pRB and an increase in the number of population doublings.
Subject(s)
Carrier Proteins/genetics , Cell Transformation, Viral/genetics , Base Sequence , Carrier Proteins/metabolism , Cell Line , Cyclin-Dependent Kinase Inhibitor p16 , Fibroblasts/metabolism , Fibroblasts/pathology , G1 Phase , Gene Expression Regulation , Genes, Tumor Suppressor , Humans , Molecular Sequence Data , RNA/genetics , Simian virus 40 , Transcription, GeneticABSTRACT
The functions of basic helix-loop-helix (bHLH) transcription factors in activating differentiation-linked gene expression and in inducing G1 cell cycle arrest are negatively regulated by members of the Id family of HLH proteins. These bHLH antagonists are induced during a mitogenic signalling response, and they function by sequestering their bHLH targets in inactive heterodimers that are unable to bind to specific gene regulatory (E box) sequences. Recently, cyclin E-Cdk2- and cyclin A-Cdk2-dependent phosphorylation of a single conserved serine residue (Ser5) in Id2 has been shown to occur during late G1-to-S phase transition of the cell cycle, and this neutralizes the function of Id2 in abrogating E-box-dependent bHLH homo- or heterodimer complex formation in vitro (E. Hara, M. Hall, and G. Peters, EMBO J. 16:332-342, 1997). We now show that an analogous cell-cycle-regulated phosphorylation of Id3 alters the specificity of Id3 for abrogating both E-box-dependent bHLH homo- or heterodimer complex formation in vitro and E-box-dependent reporter gene function in vivo. Furthermore, compared with wild-type Id3, an Id3 Asp5 mutant (mimicking phosphorylation) is unable to promote cell cycle S phase entry in transfected fibroblasts, whereas an Id3 Ala5 mutant (ablating phosphorylation) displays an activity significantly greater than that of wild-type Id3 protein. Cdk2-dependent phosphorylation therefore provides a switch during late G1-to-S phase that both nullifies an early G1 cell cycle regulatory function of Id3 and modulates its target bHLH specificity. These data also demonstrate that the ability of Id3 to promote cell cycle S phase entry is not simply a function of its ability to modulate bHLH heterodimer-dependent gene expression and establish a biologically important mechanism through which Cdk2 and Id-bHLH functions are integrated in the coordination of cell proliferation and differentiation.
Subject(s)
CDC2-CDC28 Kinases , Cell Cycle/physiology , Cyclin-Dependent Kinases/metabolism , Neoplasm Proteins , Protein Serine-Threonine Kinases/metabolism , Transcription Factors/metabolism , Animals , B-Lymphocytes/cytology , B-Lymphocytes/metabolism , Baculoviridae/genetics , Cell Line , Cyclin-Dependent Kinase 2 , Escherichia coli/genetics , Gene Expression , Helix-Loop-Helix Motifs/genetics , Helix-Loop-Helix Motifs/physiology , Humans , Inhibitor of Differentiation Proteins , Mutation , Phosphorylation , Rats , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Spodoptera , Transcription Factors/genetics , TransfectionABSTRACT
To investigate the mode of action of the p16(INK4a) tumor suppressor protein, we have established U2-OS cells in which the expression of p16(INK4a) can be regulated by addition or removal of isopropyl-beta-D-thiogalactopyranoside. As expected, induction of p16(INK4a) results in a G1 cell cycle arrest by inhibiting phosphorylation of the retinoblastoma protein (pRb) by the cyclin-dependent kinases CDK4 and CDK6. However, induction of p16(INK4a) also causes marked inhibition of CDK2 activity. In the case of cyclin E-CDK2, this is brought about by reassortment of cyclin, CDK, and CDK-inhibitor complexes, particularly those involving p27(KIP1). Size fractionation of the cellular lysates reveals that a substantial proportion of CDK4 participates in active kinase complexes of around 200 kDa. Upon induction of p16(INK4a), this complex is partly dissociated, and the majority of CDK4 is found in lower-molecular-weight fractions consistent with the formation of a binary complex with p16(INK4a). Sequestration of CDK4 by p16(INK4a) allows cyclin D1 to associate increasingly with CDK2, without affecting its interactions with the CIP/KIP inhibitors. Thus, upon the induction of p16(INK4a), p27(KIP1) appears to switch its allegiance from CDK4 to CDK2, and the accompanying reassortment of components leads to the inhibition of cyclin E-CDK2 by p27(KIP1) and p21(CIP1). Significantly, p16(INK4a) itself does not appear to form higher-order complexes, and the overwhelming majority remains either free or forms binary associations with CDK4 and CDK6.
Subject(s)
CDC2-CDC28 Kinases , Cell Cycle Proteins , Cyclin-Dependent Kinase Inhibitor p16/biosynthesis , Cyclin-Dependent Kinases/metabolism , Cyclins/metabolism , Microtubule-Associated Proteins/metabolism , Protein Serine-Threonine Kinases/metabolism , Proto-Oncogene Proteins , Tumor Suppressor Proteins , Cell Cycle , Cell Division , Cyclin-Dependent Kinase 2 , Cyclin-Dependent Kinase 4 , Cyclin-Dependent Kinase Inhibitor p16/genetics , Cyclin-Dependent Kinase Inhibitor p21 , Cyclin-Dependent Kinase Inhibitor p27 , Cyclin-Dependent Kinases/antagonists & inhibitors , Enzyme Inhibitors , Humans , Protein Serine-Threonine Kinases/antagonists & inhibitors , Time Factors , Tumor Cells, CulturedABSTRACT
BACKGROUND: Narcotics have traditionally been used to control pain after augmentation mammaplasty, but they have adverse side effects, including addiction potential, clouded sensorium, nausea, and respiratory depression. Alternative strategies for managing postoperative pain are expensive, cumbersome, and also have their own risks. While long-term use of celecoxib has been associated with an increased risk of serious adverse cardiovascular effects, no problems have been reported with short-term use. OBJECTIVE: The purpose of this study was to determine whether the addition of celecoxib, a selective nonsteroidal anti-inflammatory, to an analgesic regimen reduced narcotic use and pain following augmentation mammaplasty. METHODS: One hundred patients underwent submuscular augmentation mammaplasty with smooth saline-filled mammary prostheses using an intravenous sedation technique. Group A (N = 50) used hydrocodone to manage postoperative pain. Group B (N = 50) used celecoxib 400 mg 1 to 2 hours before surgery and then daily in addition to hydrocodone postoperatively. Narcotic use, incidence of nausea, and complications were recorded. Pain was assessed daily with a Likert pain scale from 0 (no pain) to 10 (severe pain). RESULTS: Group A used 110 +/- 34 mg hydrocodone during the immediate 7-day postoperative period and reported an average pain scale score of 5.1. Group B, which used celecoxib, used 34 +/- 22 mg hydrocodone during the same period with an average pain scale score of 3.3. These differences were statistically significant (P < .05). Group B experienced 53% less nausea than Group A. There were no significant differences between the groups regarding age, implant size, or complications. CONCLUSIONS: Perioperative celecoxib administration in patients undergoing augmentation mammaplasty significantly reduced postoperative narcotic use, pain, and nausea. Its use should facilitate the patient's ability to resume everyday activities following surgery.
ABSTRACT
The roles of two species of cytochrome P-450, the major cytochrome P-450 components of liver microsomes of phenobarbital-treated rats (PB-P-450) and 3-methylcholanthrene-treated rats (MC-P-448), were studied in the metabolism of benzo(a)pyrene in rat liver microsomes in vitro. Benzo(a)pyrene was incubated with polychlorinated biphenyl-treated rat liver microsomes, in which PB-P-450 and MC-P-448 constituted about 45 and 24% of the total cytochrome P-450, respectively. Then the metabolites were separated into those soluble in ethyl acetate and in water, and those covalently bound to protein. Using high-pressure liquid chromatography, the ethyl acetate-soluble metabolites were separated into three major groups, phenols, quinones, and dihydrodiols, including peaks of three unknown materials. Addition of anti-MC-P-448 immunoglobulin to the reaction mixture completely inhibited the formation of all ethyl acetate-soluble metabolites. In contrast, anti-PB-P-450 immunoglobulin did not inhibit the formations of 4,5-dihydro-4,5-dihydroxybenzo(a)pyrene and 3-hydroxybenzo(a)pyrene; partially inhibited the formations of 7,8-dihydro-7,8-dihydroxybenzo(a)pyrene, 9, 10-dihydro-9, 10-dihydroxybenzo(a)pyrene, and the three unknown materials; and caused 30 to 40% enhancement of the formations of 9-hydroxy-benzo(a)pyrene and benzo(a)pyrene-3,6-dione and 80% enhancement of that of benzo(a)pyrene-1,6-dione. Antibody against MC-P-448, but not against PB-P-450, also caused 75% inhibition of the formation of water-soluble metabolites and 85% inhibition of formation of benzo(a)pyrene metabolites covalently bound to protein. These results show that MC-P-448 is important in the metabolism of benzo(a)pyrene.