Discovering unknown human and mouse transcription factor binding sites and their characteristics from ChIP-seq data.

Transcription factor binding sites (TFBSs) are essential for gene regulation, but the number of known TFBSs remains limited. We aimed to discover and characterize unknown TFBSs by developing a computational pipeline for analyzing ChIP-seq (chromatin immunoprecipitation followed by sequencing) data. Applying it to the latest ENCODE ChIP-seq data for human and mouse, we found that using the irreproducible discovery rate as a quality-control criterion resulted in many experiments being unnecessarily discarded. By contrast, the number of motif occurrences in ChIP-seq peak regions provides a highly effective criterion, which is reliable even if supported by only one experimental replicate. In total, we obtained 2,058 motifs from 1,089 experiments for 354 human TFs and 163 motifs from 101 experiments for 34 mouse TFs. Among these motifs, 487 have not previously been reported. Mapping the canonical motifs to the human genome reveals a high TFBS density ±2 kb around transcription start sites (TSSs) with a peak at -50 bp. On average, a promoter contains 5.7 TFBSs. However, 70% of TFBSs are in introns (41%) and intergenic regions (29%), whereas only 12% are in promoters (-1 kb to +100 bp from TSSs). Notably, some TFs (e.g., CTCF, JUN, JUNB, and NFE2) have motifs enriched in intergenic regions, including enhancers. We inferred 142 cobinding TF pairs and 186 (including 115 completely) tethered binding TF pairs, indicating frequent interactions between TFs and a higher frequency of tethered binding than cobinding. This study provides a large number of previously undocumented motifs and insights into the biological and genomic features of TFBSs.

Origin and Evolution of Nitrogen Fixation in Prokaryotes.

The origin of nitrogen fixation is an important issue in evolutionary biology. While nitrogen is required by all living organisms, only a small fraction of bacteria and archaea can fix nitrogen. The prevailing view is that nitrogen fixation first evolved in archaea and was later transferred to bacteria. However, nitrogen-fixing (Nif) bacteria are far larger in number and far more diverse in ecological niches than Nif archaea. We, therefore, propose the bacteria-first hypothesis, which postulates that nitrogen fixation first evolved in bacteria and was later transferred to archaea. As >30,000 prokaryotic genomes have been sequenced, we conduct an in-depth comparison of the two hypotheses. We first identify the six genes involved in nitrogen fixation in all sequenced prokaryotic genomes and then reconstruct phylogenetic trees using the six Nif proteins individually or in combination. In each of these trees, the earliest lineages are bacterial Nif protein sequences and in the oldest clade (group) the archaeal sequences are all nested inside bacterial sequences, suggesting that the Nif proteins first evolved in bacteria. The bacteria-first hypothesis is further supported by the observation that the majority of Nif archaea carry the major bacterial Mo (molybdenum) transporter (ModABC) rather than the archaeal Mo transporter (WtpABC). Moreover, in our phylogeny of all available ModA and WtpA protein sequences, the earliest lineages are bacterial sequences while archaeal sequences are nested inside bacterial sequences. Furthermore, the bacteria-first hypothesis is supported by available isotopic data. In conclusion, our study strongly supports the bacteria-first hypothesis.

Maize ANT1 modulates vascular development, chloroplast development, photosynthesis, and plant growth.

Arabidopsis AINTEGUMENTA (ANT), an AP2 transcription factor, is known to control plant growth and floral organogenesis. In this study, our transcriptome analysis and in situ hybridization assays of maize embryonic leaves suggested that maize ANT1 (ZmANT1) regulates vascular development. To better understand ANT1 functions, we determined the binding motif of ZmANT1 and then showed that ZmANT1 binds the promoters of millet SCR1, GNC, and AN3, which are key regulators of Kranz anatomy, chloroplast development, and plant growth, respectively. We generated a mutant with a single-codon deletion and two frameshift mutants of the ANT1 ortholog in the C4 millet Setaria viridis by the CRISPR/Cas9 technique. The two frameshift mutants displayed reduced photosynthesis efficiency and growth rate, smaller leaves, and lower grain yields than wild-type (WT) plants. Moreover, their leaves sporadically exhibited distorted Kranz anatomy and vein spacing. Conducting transcriptomic analysis of developing leaves in the WT and the three mutants we identified differentially expressed genes (DEGs) in the two frameshift mutant lines and found many down-regulated DEGs enriched in photosynthesis, heme, tetrapyrrole binding, and antioxidant activity. In addition, we predicted many target genes of ZmANT1 and chose 13 of them to confirm binding of ZmANT1 to their promoters. Based on the above observations, we proposed a model for ANT1 regulation of cell proliferation and leaf growth, vascular and vein development, chloroplast development, and photosynthesis through its target genes. Our study revealed biological roles of ANT1 in several developmental processes beyond its known roles in plant growth and floral organogenesis.

Identifying Primate ACE2 Variants That Confer Resistance to SARS-CoV-2.

SARS-CoV-2 infects humans through the binding of viral S-protein (spike protein) to human angiotensin I converting enzyme 2 (ACE2). The structure of the ACE2-S-protein complex has been deciphered and we focused on the 27 ACE2 residues that bind to S-protein. From human sequence databases, we identified nine ACE2 variants at ACE2-S-protein binding sites. We used both experimental assays and protein structure analysis to evaluate the effect of each variant on the binding affinity of ACE2 to S-protein. We found one variant causing complete binding disruption, two and three variants, respectively, strongly and mildly reducing the binding affinity, and two variants strongly enhancing the binding affinity. We then collected the ACE2 gene sequences from 57 nonhuman primates. Among the 6 apes and 20 Old World monkeys (OWMs) studied, we found no new variants. In contrast, all 11 New World monkeys (NWMs) studied share four variants each causing a strong reduction in binding affinity, the Philippine tarsier also possesses three such variants, and 18 of the 19 prosimian species studied share one variant causing a strong reduction in binding affinity. Moreover, one OWM and three prosimian variants increased binding affinity by >50%. Based on these findings, we proposed that the common ancestor of primates was strongly resistant to and that of NWMs was completely resistant to SARS-CoV-2 and so is the Philippine tarsier, whereas apes and OWMs, like most humans, are susceptible. This study increases our understanding of the differences in susceptibility to SARS-CoV-2 infection among primates.

Many human RNA viruses show extraordinarily stringent selective constraints on protein evolution.

How negative selection, positive selection, and population size contribute to the large variation in nucleotide substitution rates among RNA viruses remains unclear. Here, we studied the ratios of nonsynonymous-to-synonymous substitution rates (dN/dS) in protein-coding genes of human RNA and DNA viruses and mammals. Among the 21 RNA viruses studied, 18 showed a genome-average dN/dS from 0.01 to 0.10, indicating that over 90% of nonsynonymous mutations are eliminated by negative selection. Only HIV-1 showed a dN/dS (0.31) higher than that (0.22) in mammalian genes. By comparing the dN/dS values among genes in the same genome and among species or strains, we found that both positive selection and population size play significant roles in the dN/dS variation among genes and species. Indeed, even in flaviviruses and picornaviruses, which showed the lowest ratios among the 21 species studied, positive selection appears to have contributed significantly to dN/dS We found the view that positive selection occurs much more frequently in influenza A subtype H3N2 than subtype H1N1 holds only for the hemagglutinin and neuraminidase genes, but not for other genes. Moreover, we found no support for the view that vector-borne RNA viruses have lower dN/dS ratios than non-vector-borne viruses. In addition, we found a correlation between dN and dS, implying a correlation between dN and the mutation rate. Interestingly, only 2 of the 8 DNA viruses studied showed a dN/dS < 0.10, while 4 showed a dN/dS > 0.22. These observations increase our understanding of the mechanisms of RNA virus evolution.

Comparative transcriptomics method to infer gene coexpression networks and its applications to maize and rice leaf transcriptomes.

Time-series transcriptomes of a biological process obtained under different conditions are useful for identifying the regulators of the process and their regulatory networks. However, such data are 3D (gene expression, time, and condition), and there is currently no method that can deal with their full complexity. Here, we developed a method that avoids time-point alignment and normalization between conditions. We applied it to analyze time-series transcriptomes of developing maize leaves under light-dark cycles and under total darkness and obtained eight time-ordered gene coexpression networks (TO-GCNs), which can be used to predict upstream regulators of any genes in the GCNs. One of the eight TO-GCNs is light-independent and likely includes all genes involved in the development of Kranz anatomy, which is a structure crucial for the high efficiency of photosynthesis in C4 plants. Using this TO-GCN, we predicted and experimentally validated a regulatory cascade upstream of SHORTROOT1, a key Kranz anatomy regulator. Moreover, we applied the method to compare transcriptomes from maize and rice leaf segments and identified regulators of maize C4 enzyme genes and RUBISCO SMALL SUBUNIT2 Our study provides not only a powerful method but also novel insights into the regulatory networks underlying Kranz anatomy development and C4 photosynthesis.

The Cone Opsin Repertoire of Osteoglossomorph Fishes: Gene Loss in Mormyrid Electric Fish and a Long Wavelength-Sensitive Cone Opsin That Survived 3R.

Vertebrates have four classes of cone opsin genes derived from two rounds of genome duplication. These are short wavelength sensitive 1(SWS1), short wavelength sensitive 2(SWS2), medium wavelength sensitive (RH2), and long wavelength sensitive (LWS). Teleosts had another genome duplication at their origin and it is believed that only one of each cone opsin survived the ancestral teleost duplication event. We tested this by examining the retinal cones of a basal teleost group, the osteoglossomorphs. Surprisingly, this lineage has lost the typical vertebrate green-sensitive RH2 opsin gene and, instead, has a duplicate of the LWS opsin that is green sensitive. This parallels the situation in mammalian evolution in which the RH2 opsin gene was lost in basal mammals and a green-sensitive opsin re-evolved in Old World, and independently in some New World, primates from an LWS opsin gene. Another group of fish, the characins, possess green-sensitive LWS cones. Phylogenetic analysis shows that the evolution of green-sensitive LWS opsins in these two teleost groups derives from a common ancestral LWS opsin that acquired green sensitivity. Additionally, the nocturnally active African weakly electric fish (Mormyroideae), which are osteoglossomorphs, show a loss of the SWS1 opsin gene. In comparison with the independently evolved nocturnally active South American weakly electric fish (Gymnotiformes) with a functionally monochromatic LWS opsin cone retina, the presence of SWS2, LWS, and LWS2 cone opsins in mormyrids suggests the possibility of color vision.

Transcriptome dynamics of developing maize leaves and genomewide prediction of cis elements and their cognate transcription factors.

Maize is a major crop and a model plant for studying C4 photosynthesis and leaf development. However, a genomewide regulatory network of leaf development is not yet available. This knowledge is useful for developing C3 crops to perform C4 photosynthesis for enhanced yields. Here, using 22 transcriptomes of developing maize leaves from dry seeds to 192 h post imbibition, we studied gene up- and down-regulation and functional transition during leaf development and inferred sets of strongly coexpressed genes. More significantly, we developed a method to predict transcription factor binding sites (TFBSs) and their cognate transcription factors (TFs) using genomic sequence and transcriptomic data. The method requires not only evolutionary conservation of candidate TFBSs and sets of strongly coexpressed genes but also that the genes in a gene set share the same Gene Ontology term so that they are involved in the same biological function. In addition, we developed another method to predict maize TF-TFBS pairs using known TF-TFBS pairs in Arabidopsis or rice. From these efforts, we predicted 1,340 novel TFBSs and 253 new TF-TFBS pairs in the maize genome, far exceeding the 30 TF-TFBS pairs currently known in maize. In most cases studied by both methods, the two methods gave similar predictions. In vitro tests of 12 predicted TF-TFBS interactions showed that our methods perform well. Our study has significantly expanded our knowledge on the regulatory network involved in maize leaf development.

Regulatory Divergence among Beta-Keratin Genes during Bird Evolution.

Feathers, which are mainly composed of α- and ß-keratins, are highly diversified, largely owing to duplication and diversification of ß-keratin genes during bird evolution. However, little is known about the regulatory changes that contributed to the expressional diversification of ß-keratin genes. To address this issue, we studied transcriptomes from five different parts of chicken contour and flight feathers. From these transcriptomes we inferred ß-keratin enriched co-expression modules of genes and predicted transcription factors (TFs) of ß-keratin genes. In total, we predicted 262 TF-target gene relationships in which 56 TFs regulate 91 ß-keratin genes; we validated 14 of them by in vitro tests. A dual criterion of TF enrichment and "TF-target gene" expression correlation identified 26 TFs as the major regulators of ß-keratin genes. According to our predictions, the ancestral scale and claw ß-keratin genes have common and unique regulators, whereas most feather ß-keratin genes show chromosome-wise regulation, distinct from scale and claw ß-keratin genes. Thus, after expansion from the ß-keratin gene on Chr7 to other chromosomes, which still shares a TF with scale and claw ß-keratin genes, most feather ß-keratin genes have recruited distinct or chromosome-specific regulators. Moreover, our data showed correlated gene expression profiles, positive or negative, between predicted TFs and their target genes over the five studied feather regions. Therefore, regulatory divergences among feather ß-keratin genes have contributed to structural differences among different parts of feathers. Our study sheds light on how feather ß-keratin genes have diverged in regulation from scale and claw ß-keratin genes and among themselves.

Maize and millet transcription factors annotated using comparative genomic and transcriptomic data.

BACKGROUND: Transcription factors (TFs) contain DNA-binding domains (DBDs) and regulate gene expression by binding to specific DNA sequences. In addition, there are proteins, called transcription coregulators (TCs), which lack DBDs but can alter gene expression through interaction with TFs or RNA Polymerase II. Therefore, it is interesting to identify and classify the TFs and TCs in a genome. In this study, maize (Zea mays) and foxtail millet (Setaria italica), two important species for the study of C4 photosynthesis and kranz anatomy, were selected. RESULT: We conducted a comprehensive genome-wide annotation of TFs and TCs in maize B73 and in two strains of foxtail millet, Zhang gu and Yugu1, and classified them into families. To gain additional support for our predictions, we searched for their homologous genes in Arabidopsis or rice and studied their gene expression level using RNA-seq and microarray data. We identified many new TF and TC families in these two species, and described some evolutionary and functional aspects of the 9 new maize TF families. Moreover, we detected many pseudogenes and transposable elements in current databases. In addition, we examined tissue expression preferences of TF and TC families and identified tissue/condition-specific TFs and TCs in maize and millet. Finally, we identified potential C4-related TF and TC genes in maize and millet. CONCLUSIONS: Our results significantly expand current TF and TC annotations in maize and millet. We provided supporting evidence for our annotation from genomic and gene expression data and identified TF and TC genes with tissue preference in expression. Our study may facilitate the study of regulation of gene expression, tissue morphogenesis, and C4 photosynthesis in maize and millet. The data we generated in this study are available at http://sites.google.com/site/jjlmmtf.

High short-term and long-term excess mortality in geriatric patients after hip fracture: a prospective cohort study in Taiwan.

BACKGROUND: Hip fracture has a high mortality rate, but the actual level of long-term excess mortality and its impact on population-wide mortality remains controversial. The present prospective study investigated short- and long-term excess mortality after hip fractures with adjustment of other risk factors. We calculated the population attributable risk proportion (PARP) to assess the impact of each risk factor on excess mortality. METHODS: We recruited 217 elders with hip fractures and 215 age- and sex-matched patients without fractures from the geriatric department of the same hospital. The mean follow-up time was 46.1 months (range: 35 to 57 months). We recorded data on 55 covariates, including baseline details about health, function, and bone mineral density. We used the multivariate Cox proportional hazards model to analyze hazard ratios (HRs) of short-term (<12 months follow-up) and long-term (≧ 2 months follow-up) excess mortality for each covariate and calculated their PARP. RESULTS: Patients with hip fractures had a higher short-term mortality than non-fractured patients, and the long-term excess mortality associated with hip fracture remained high. The significant risk factors for short-term mortality were hip fracture, comorbidities, and lower (below cutoff) Mini Mental State Examination score with HRs of 2.4, 2.3, and 2.3, respectively. Their PARPs were 44.7%, 38.1%, and 34.3%, respectively. The significant risk factors for long-term mortality were hip fracture (HR: 2.7; PARP: 48.0%), lower T-score (HR: 3.3; PARP: 36.2%), lower body mass index (HR: 2.5; PARP: 42.8%), comorbidities (HR: 2.1; PARP: 34.8%), difficulty in activities of daily living (HR: 1.9; PARP: 31.8%), and smoking (HR: 2.5; PARP: 19.2%). CONCLUSIONS: After comprehensive adjustment, hip fracture was a significant risk factor and contributed the most to long-term as well as short-term excess mortality. Its adequate prevention and treatment should be targeted.

The genomic evolution of visual opsin genes in amphibians.

Among tetrapod (terrestrial) vertebrates, amphibians remain more closely tied to an amphibious lifestyle than amniotes, and their visual opsin genes may be adapted to this lifestyle. Previous studies have discussed physiological, morphological, and molecular changes in the evolution of amphibian vision. We predicted the locations of the visual opsin genes, their neighboring genes, and the tuning sites of the visual opsins, in 39 amphibian genomes. We found that all of the examined genomes lacked the Rh2 gene. The caecilian genomes have further lost the SWS1 and SWS2 genes; only the Rh1 and LWS genes were retained. The loss of the SWS1 and SWS2 genes in caecilians may be correlated with their cryptic lifestyles. The opsin gene syntenies were predicted to be highly similar to those of other bony vertebrates. Moreover, dual syntenies were identified in allotetraploid Xenopus laevis and X. borealis. Tuning site analysis showed that only some Caudata species might have UV vision. In addition, the S164A that occurred several times in LWS evolution might either functionally compensate for the Rh2 gene loss or fine-tuning visual adaptation. Our study provides the first genomic evidence for a caecilian LWS gene and a genomic viewpoint of visual opsin genes by reviewing the gains and losses of visual opsin genes, the rearrangement of syntenies, and the alteration of spectral tuning in the course of amphibians' evolution.

Conserved regulatory switches for the transition from natal down to juvenile feather in birds.

The transition from natal downs for heat conservation to juvenile feathers for simple flight is a remarkable environmental adaptation process in avian evolution. However, the underlying epigenetic mechanism for this primary feather transition is mostly unknown. Here we conducted time-ordered gene co-expression network construction, epigenetic analysis, and functional perturbations in developing feather follicles to elucidate four downy-juvenile feather transition events. We report that extracellular matrix reorganization leads to peripheral pulp formation, which mediates epithelial-mesenchymal interactions for branching morphogenesis. α-SMA (ACTA2) compartmentalizes dermal papilla stem cells for feather renewal cycling. LEF1 works as a key hub of Wnt signaling to build rachis and converts radial downy to bilateral symmetry. Novel usage of scale keratins strengthens feather sheath with SOX14 as the epigenetic regulator. We show that this primary feather transition is largely conserved in chicken (precocial) and zebra finch (altricial) and discuss the possibility that this evolutionary adaptation process started in feathered dinosaurs.

Hip fracture risk assessment: artificial neural network outperforms conditional logistic regression in an age- and sex-matched case control study.

BACKGROUND: Osteoporotic hip fractures with a significant morbidity and excess mortality among the elderly have imposed huge health and economic burdens on societies worldwide. In this age- and sex-matched case control study, we examined the risk factors of hip fractures and assessed the fracture risk by conditional logistic regression (CLR) and ensemble artificial neural network (ANN). The performances of these two classifiers were compared. METHODS: The study population consisted of 217 pairs (149 women and 68 men) of fractures and controls with an age older than 60 years. All the participants were interviewed with the same standardized questionnaire including questions on 66 risk factors in 12 categories. Univariate CLR analysis was initially conducted to examine the unadjusted odds ratio of all potential risk factors. The significant risk factors were then tested by multivariate analyses. For fracture risk assessment, the participants were randomly divided into modeling and testing datasets for 10-fold cross validation analyses. The predicting models built by CLR and ANN in modeling datasets were applied to testing datasets for generalization study. The performances, including discrimination and calibration, were compared with non-parametric Wilcoxon tests. RESULTS: In univariate CLR analyses, 16 variables achieved significant level, and six of them remained significant in multivariate analyses, including low T score, low BMI, low MMSE score, milk intake, walking difficulty, and significant fall at home. For discrimination, ANN outperformed CLR in both 16- and 6-variable analyses in modeling and testing datasets (p?

Effects of merged holes, partial thread removal, and offset holes on fatigue strengths of titanium locking plates.

BACKGROUND: This study investigated the effects of screw hole merging, thread removal, and screw hole offset on the mechanical properties of locking plates. METHODS: Finite element models were used to develop the optimal design of the merged holes. Four titanium locking plates with different hole designs were analyzed. Type I had threaded round holes. Type II had merged holes. Type III had merged holes with partial thread removal. Type IV had threaded offset holes. Mechanical experiments similar to finite element analyses were conducted and compared. Screw bending tests were used to assess the screw holding power. FINDINGS: Finite element analyses showed the optimal merging distance between two round screw holes was 3.5 mm with 2/3 circumferences in each hole. The stresses of types II and III were respectively 6.42% and 7.33%, lower than that of type I. The stress of type IV was 1.66% higher than that of type I. In the mechanical tests, the fatigue lives of types II and III were respectively 3.86 and 7.16 times higher than that of type I. The fatigue life of type IV was 37% lower than that of type I. The differences in the bending strengths of screws were insignificant. INTERPRETATION: Merging holes could mitigate screw hole stress and increase the fatigue lives of the plates significantly. Partial thread removal could further improve the fatigue life. Merging holes and thread removal did not decrease the screw holding power significantly. The fatigue lives were significantly decreased in plates with offset holes.

Risk factors for hip fracture sites and mortality in older adults.

BACKGROUND: Our study was designed to (1) investigate the risk factors associated with cervical or trochanteric hip fractures; and (2) identify the risk factors for increased mortality in the elderly population sustaining hip fractures, after adjusting the miscellaneous baseline prefracture conditions. METHODS: Two hundred seventeen elder patients with first-time, low-trauma hip fractures were enrolled. The follow-up time ranged from 35 months to 56 months. Potential risk factors for hip fracture types included (1) the 77 items on the self-reported questionnaire; (2) the body height, weight, and body mass index; (3) tests of coordination, handgrip strength, and peak expiratory flow rate; and (4) the bone mineral density variables. GTFN ratio was defined as the bone mineral density ratio between the greater trochanter and the femoral neck. Multivariate logistic regression and Cox regression models were used for analysis. The population attributable risk proportion of death to each significant factor was estimated. RESULTS: Risk factors for trochanteric fractures include a GTFN ratio ≤0.81, being male, and an age >80 years. Risk factors for higher mortality after hip fracture included trochanteric fracture, body mass index ≤20 (kg/m), poor self-assessed health status, peak expiratory flow rate ≤215 (L/min), being male, illiteracy, and coordination abnormality, in the declining order of population attributable risk proportion. Trochanteric fractures had a significantly higher cumulative mortality at 36 months, 48 months, and 60 months than cervical fractures. CONCLUSIONS: The novel GTFN ratio was associated with hip fracture sites. Clinically, cervical and trochanteric fractures represent different disease entities because of the difference in their mortality rates.

A neurogenetic approach to a multiobjective design optimization of spinal pedicle screws.

A pedicle screw fixation has been widely used to treat spinal diseases. Clinical reports have shown that the weakest part of the spinal fixator is the pedicle screw. However, previous studies have only focused on either screw breakage or screw loosening. There have been no studies that have addressed the multiobjective design optimization of the pedicle screws. The multiobjective optimization methodology was applied and it consisted of finite element method, Taguchi method, artificial neural networks, and genetic algorithms. Three-dimensional finite element models for both the bending strength and the pullout strength of the pedicle screw were first developed and arranged on an L(25) orthogonal array. Then, artificial neural networks were used to create two objective functions. Finally, the optimum solutions of the pedicle screws were obtained by genetic algorithms. The results showed that the optimum designs had higher bending and pullout strengths compared with commercially available screws. The optimum designs of pedicle screw revealed excellent biomechanical performances. The neurogenetic approach has effectively decreased the time and effort required for searching for the optimal designs of pedicle screws and has directly provided the selection information to surgeons.

Allografting in locked nailing and interfragmentary wiring for humeral nonunions.

UNLABELLED: In this prospective study, we compared outcomes after repair of humeral nonunions when morsellized fresh-frozen allograft or autograft was used to augment repair by intramedullary nailing. Sixty-five patients with humeral shaft nonunions of greater than 6 months' duration and gross instability at the nonunion site were included and treated by locked nailing, interfragmentary wiring, and bone grafting. Graft type was determined by patient preference. Outcomes assessed included union rate and functional recovery of the arm. Secondary end points included operative blood loss, operation time, hospital stay, time to fracture healing, and complications. Twenty-eight patients with autografts and 36 with allografts were followed up more than 2 years. The baseline conditions of the two groups were similar. The autograft group had greater blood loss and longer operative time than the allograft group. The autograft group also had a longer hospital stay. The healing rate, time to healing, and functional scores did not differ between these two groups. In the autograft group, 43% reported pain and limited mobility as a result of the donor site. We concluded that when used in association with locked nailing for humeral nonunions, allografts can achieve treatment results similar to autografts but without donor site complications. LEVEL OF EVIDENCE: Level III, prognostic study. See Guidelines for Authors for a complete description of levels of evidence.

Half-threaded holes markedly increase the fatigue life of locking plates without compromising screw stability.

AIMS: To determine whether half-threaded screw holes in a new titanium locking plate design can substantially decrease the notch effects of the threads and increase the plate fatigue life. METHODS: Three types (I to III) of titanium locking plates were fabricated to simulate plates used in the femur, tibia, and forearm. Two copies of each were fabricated using full- and half-threaded screw holes (called A and B, respectively). The mechanical strengths of the plates were evaluated according to the American Society for Testing and Materials (ASTM) F382-14, and the screw stability was assessed by measuring the screw removal torque and bending strength. RESULTS: The B plates had fatigue lives 11- to 16-times higher than those of the A plates. Before cyclic loading, the screw removal torques were all higher than the insertion torques. However, after cyclic loading, the removal torques were similar to or slightly lower than the insertion torques (0% to 17.3%), although those of the B plates were higher than those of the A plates for all except the type III plates (101%, 109.8%, and 93.8% for types I, II, and III, respectively). The bending strengths of the screws were not significantly different between the A and B plates for any of the types. CONCLUSION: Removing half of the threads from the screw holes markedly increased the fatigue life of the locking plates while preserving the tightness of the screw heads and the bending strength of the locking screws. However, future work is necessary to determine the relationship between the notch sensitivity properties and titanium plate design.Cite this article: Bone Joint Res 2020;9(10):645-652.

Contradictory working length effects in locked plating of the distal and middle femoral fractures-a biomechanical study.

BACKGROUND: Working length have been reported to affect the plate stress and fixation stiffness. However, the results of previous studies have been controversial. The present study was to determine working length effects on different locations of femoral bone gap. METHODS: Five composite femurs with wide bone gaps at five levels (G1, 2, 3, 5, and 7), were fixed with locking plates. G1-3, G5 and G7 represented gaps at distal femur, distal-middle femur and middle femur respectively. Strain gauges were applied near the screw holes. The plate-bone constructs were loaded through a hemicylinder on the femoral head with total constraints at the distal femur. The micro-strains, axial stiffness and interfragmentary motions were recorded. Then the locking screws were removed one by one and the tests were re-run. The working length effects were compared and correlated. FINDINGS: In distal femurs (G1-3), long working length was negatively correlated with the highest strains (r = -0.97, -0.95 and - 0.95, p < 0.01) and axial stiffness (r = -1, -0.96 and -0.99, p < 0.01). In distal-middle femurs (G5), as the working length increased, the highest strain decreased initially and then increased (r = 0.81, p = 0.026) and the axial stiffness decreased (r = -0.98, p < 0.01). In middle femurs (G7), the highest strain and gap motions were much higher than that in the other groups and not significantly correlated with the working length change. INTERPRETATION: Long working length could reduce the highest plate strain in distal femurs, but had no significant effects in middle femurs. The working length effects were markedly affected by the loading and boundary conditions.