A systematic review of short metaphyseal loading cementless stems in hip arthroplasty.

AIMS: Short-stemmed femoral implants have been used for total hip arthroplasty (THA) in young and active patients to conserve bone, provide physiological loading, and reduce the incidence of thigh pain. Only short- to mid-term results have been presented and there have been concerns regarding component malalignment, incorrect sizing, and subsidence. This systematic review reports clinical and radiological outcomes, complications, revision rates, and implant survival in THA using short-stemmed femoral components. MATERIALS AND METHODS: A literature review was performed using the EMBASE, Medline, and Cochrane databases. Strict inclusion and exclusion criteria were used to identify studies reporting clinical and radiological follow-up for short-stemmed hip arthroplasties. RESULTS: A total of 28 studies were eligible for inclusion. This included 5322 hips in 4657 patients with a mean age of 59 years (13 to 94). The mean follow-up was 6.1 years (0.5 to 20). The mean Harris Hip Score improved from 46 (0 to 100) to 92 (39 to 100). The mean Oxford Hip Score improved from 25 (2 to 42.5) to 35 (12.4 to 48). The mean Western Ontario & McMaster Universities Osteoarthritis Index improved from 54 (2 to 95) to 22 (0 to 98). Components were aligned in a neutral coronal alignment in up to 90.9% of cases. A total of 15 studies reported component survivorship, which was 98.6% (92% to 100%) at a mean follow-up of 12.1 years. CONCLUSION: Short-stemmed femoral implants show similar improvement in clinical and radiological outcomes compared with conventional length implants. Only mid-term survivorship, however, is known. An abundance of short components have been developed and used commercially without staged clinical trials. Long-term survival is still unknown for many of these components. There remains tension between innovation and the moral duty to ensure that the introduction of new implants is controlled until safety and patient benefit are demonstrated. Implant innovation and subsequent use should be driven by proven clinical outcomes, rather than market and financial forces, and ethical practice must be ensured. Cite this article: Bone Joint J 2019;101-B:502-511.

Characterization of Wnt gene expression in developing and postnatal hair follicles and identification of Wnt5a as a target of Sonic hedgehog in hair follicle morphogenesis.

Mutations in WNT effector genes perturb hair follicle morphogenesis, suggesting key roles for WNT proteins in this process. We show that expression of Wnts 10b and 10a is upregulated in placodes at the onset of follicle morphogenesis and in postnatal hair follicles beginning a new cycle of hair growth. The expression of additional Wnt genes is observed in follicles at later stages of differentiation. Among these, we find that Wnt5a is expressed in the developing dermal condensate of wild type but not Sonic hedgehog (Shh)-null embryos, indicating that Wnt5a is a target of SHH in hair follicle morphogenesis. These results identify candidates for several key follicular signals and suggest that WNT and SHH signaling pathways interact to regulate hair follicle morphogenesis.

Members of the bHLH-PAS family regulate Shh transcription in forebrain regions of the mouse CNS.

The secreted protein sonic hedgehog (Shh) is required to establish patterns of cellular growth and differentiation within ventral regions of the developing CNS. The expression of Shh in the two tissue sources responsible for this activity, the axial mesoderm and the ventral midline of the neural tube, is regulated along the anteroposterior neuraxis. Separate cis-acting regulatory sequences have been identified which direct Shh expression to distinct regions of the neural tube, supporting the view that multiple genes are involved in activating Shh transcription along the length of the CNS. We show here that the activity of one Shh enhancer, which directs reporter expression to portions of the ventral midbrain and diencephalon, overlaps both temporally and spatially with the expression of Sim2. Sim2 encodes a basic helix-loop-helix (bHLH-PAS) PAS domain containing transcriptional regulator whose Drosophila homolog, single-minded, is a master regulator of ventral midline development. Both vertebrate and invertebrate Sim family members were found sufficient for the activation of the Shh reporter as well as endogenous Shh mRNA. Although Shh expression is maintained in Sim2(-)(/)(-) embryos, it was determined to be absent from the rostral midbrain and caudal diencephalon of embryos carrying a dominant-negative transgene that disrupts the function of bHLH-PAS proteins. Together, these results suggest that bHLH-PAS family members are required for the regulation of Shh transcription within aspects of the ventral midbrain and diencephalon.

A role for Gbx2 in repression of Otx2 and positioning the mid/hindbrain organizer.

The mid/hindbrain (MHB) junction can act as an organizer to direct the development of the midbrain and anterior hindbrain. In mice, Otx2 is expressed in the forebrain and midbrain and Gbx2 is expressed in the anterior hindbrain, with a shared border at the level of the MHB organizer. Here we show that, in Gbx2-/- mutants, the earliest phenotype is a posterior expansion of the Otx2 domain during early somite stages. Furthermore, organizer genes are expressed at the shifted Otx2 border, but not in a normal spatial relationship. To test whether Gbx2 is sufficient to position the MHB organizer, we transiently expressed Gbx2 in the caudal Otx2 domain and found that the Otx2 caudal border was indeed shifted rostrally and a normal appearing organizer formed at this new Otx2 border. Transgenic embryos then showed an expanded hindbrain and a reduced midbrain at embryonic day 9.5-10. We propose that formation of a normal MHB organizer depends on a sharp Otx2 caudal border and that Gbx2 is required to position and sharpen this border.

Regionalization of Sonic hedgehog transcription along the anteroposterior axis of the mouse central nervous system is regulated by Hnf3-dependent and -independent mechanisms.

The axial midline mesoderm and the ventral midline of the neural tube, the floor plate, share the property of being a source of the secreted protein, Sonic hedgehog (Shh), which has the capacity to induce a variety of ventral cell types along the length of the mouse CNS. To gain insight into the mechanisms by which Shh transcription is initiated in these tissues, we set out to identify the cis-acting sequences regulating Shh gene expression. As an approach, we have tested genomic clones encompassing 35 kb of the Shh locus for their ability to direct a lacZ reporter gene to the temporally and spatially restricted confines of the Shh expression domains in transgenic mice. Three enhancers were identified that directed lacZ expression to distinct regions along the anteroposterior axis including the ventral midline of the spinal cord, hindbrain, rostral midbrain and caudal diencephalon, suggesting that multiple transcriptional regulators are required to initiate Shh gene expression within the CNS. In addition, regulatory sequences were also identified that directed reporter expression to the notochord, albeit, under limited circumstances. Sequence analysis of the genomic clones responsible for enhancer activity from a variety of organisms, including mouse, chicken and human, have identified highly conserved binding sites for the hepatocyte nuclear factor 3 (Hnf3) family of transcriptional regulators in some, but not all, of the enhancers. Moreover, the generation of mutations in the Hnf3-binding sites showed their requirement in certain, but not all, aspects of Shh reporter expression. Taken together, our results support the existence of Hnf3-dependent and -independent mechanisms in the direct activation of Shh transcription within the CNS and axial mesoderm.

Gli2 is required for induction of floor plate and adjacent cells, but not most ventral neurons in the mouse central nervous system.

Induction of the floor plate at the ventral midline of the neural tube is one of the earliest events in the establishment of dorsoventral (d/v) polarity in the vertebrate central nervous system (CNS). The secreted molecule, Sonic hedgehog, has been shown to be both necessary and sufficient for this induction. In vertebrates, several downstream components of this signalling pathway have been identified, including members of the Gli transcription factor family. In this study, we have examined d/v patterning of the CNS in Gli2 mouse mutants. We have found that the floor plate throughout the midbrain, hindbrain and spinal cord does not form in Gli2 homozygotes. Despite this, motoneurons and ventral interneurons form in their normal d/v positions at 9.5 to 12.5 days postcoitum (dpc). However, cells that are generated in the region flanking the floor plate, including dopaminergic and serotonergic neurons, were greatly reduced in number or absent in Gli2 homozygous embryos. These results suggest that early signals derived from the notochord can be sufficient for establishing the basic d/v domains of cell differentiation in the ventral spinal cord and hindbrain. Interestingly, the notochord in Gli2 mutants does not regress ventrally after 10.5 dpc, as in normal embryos. Finally, the spinal cord of Gli1/Gli2 zinc-finger-deletion double homozygous mutants appeared similar to Gli2 homozygotes, indicating that neither gene is required downstream of Shh for the early development of ventral cell fates outside the ventral midline.

Antagonizing cAMP-dependent protein kinase A in the dorsal CNS activates a conserved Sonic hedgehog signaling pathway.

Hedgehog (Hh) signaling plays a significant role in defining the polarity of a variety of tissue types along the anterior/posterior and dorsal/ventral axes in both vertebrate and invertebrate organisms. The pathway through which Hh transduces its signal is still obscure, however, recent data have implicated the cyclic AMP-dependent protein kinase A as a negative regulator of the Hh signal transduction pathway. One of the vertebrate Hh family members, Sonic hedgehog (Shh), can induce ventral neural cell types both in vivo and in vitro; high concentrations induce floor plate and lower concentrations motor neurons. To investigate whether PKA plays an active role in the suppression of ventral neural differentiation, we generated transgenic embryos expressing a dominant negative form of PKA (dnPKA) in primarily dorsal aspects of the mouse CNS. Similar to our earlier results with Shh, we observed the induction of floor plate and motor neuron markers in embryos expressing the dominant negative PKA transgene and the loss of dorsal gene expression at rostral levels. Thus suppression of PKA activity is sufficient to activate targets of the Shh signaling pathway in the vertebrate CNS suggesting that induction of ventral cell types occurs via the antagonistic action of Shh on PKA activity. Two mammalian target genes that are strongly expressed in ectopic dorsal locations in response to dnPKA are Ptc and Gli. As both of these are targets of Drosophila Hh signaling, our data point to an evolutionary conservation in both the mechanisms of signaling and the effectors of the signaling pathway.

Cloning, expression, and chromosomal location of SHH and IHH: two human homologues of the Drosophila segment polarity gene hedgehog.

The hedgehog genes encode signaling molecules that play a role in regulating embryonic morphogenesis. We have cloned and sequenced human cDNA copies of two of these genes, SHH and IHH. The SHH clone includes the full coding sequence and encodes a protein 92.4% identical to its murine homologue. The IHH clone is 89% complete and encodes a protein 94.6% identical to its murine homologue. IHH is expressed in adult kidney and liver. SHH expression was not detected in adult tissues examined; however, it is expressed in fetal intestine, liver, lung, and kidney. SHH mapped to chromosome 7q and IHH to chromosome 2 by PCR with DNA from a panel of rodent-human somatic cell hybrids. To identify the chromosomal location of SHH more precisely, a P1 genomic clone of SHH was isolated. This phage contained a CA repeat sequence tagged site that was used to map SHH relative to a polysyndactyly disease locus, using DNA prepared from affected and unaffected members of a large pedigree. SHH is closely linked, but distinct from the polysyndactyly disease locus at 7q36 (maximum lod score = 4.82, theta = 0.05) tightly linked to the EN2 locus. The murine homologues Shh, Ihh, and Dhh were mapped using (C57BL/6J x Mus spretus)F1 x C57BL/6J interspecific backcross. Shh mapped to a position 0.6 cM distal to En2 and 1.9 cM proximal to Il6 on mouse chromosome 5. This location is closely linked but distinct from the murine limb mutation Hx and syntenic to human chromosome 7q36.

Characterization of a region-specific library of microclones in the vicinity of the Bcg and splotch loci on mouse chromosome 1.

The proximal portion of mouse chromosome 1 harbors a variety of mutant loci that have yet to be characterized at the molecular level. We have constructed a library of genomic DNA fragments from the proximal portion of mouse chromosome 1 by microdissection and microcloning techniques, with the aim of generating genetic markers in close proximity to some of these mutant loci. To facilitate the genetic mapping of 27 microclones from this library, we divided a 56-cM segment of chromosome 1 between the Col3a1 and Ren 1,2 genes into eight intervals defined by anchor loci. Restriction fragment length polymorphisms were determined for each of the microclones and their segregation with the anchor loci was followed in informative animals from a panel of 252 interspecific backcross mice (C57BL/6J x Mus spretus) x C57BL/6J. We were able to assign 26 of 27 (96%) randomly selected microclones to each of the defined chromosome 1 intervals. A total of eight microclones mapped within the large interstitial deletion found in the Spr mouse mutant. Two of these clones were found to be tightly linked to the host resistance locus Bcg and at least one was found to be linked to the neural tube defect mutant splotch. Other clones mapped to intervals containing several other mouse mutants. These novel DNA markers should aid in positional cloning strategies presently employed to identify these mutant loci. These clones should also be useful in the creation of both physical and YAC contiguous maps of the proximal portion of mouse chromosome 1.

Sonic hedgehog, a member of a family of putative signaling molecules, is implicated in the regulation of CNS polarity.

We have identified three members of a mouse gene family related to the Drosophila segment polarity gene, hedgehog (hh). Like hh, they encode putative secreted proteins and are thus implicated in cell-cell interactions. One of these, Sonic hh (Shh), is expressed in the notochord, the floor plate, and the zone of polarizing activity, signaling centers that are thought to mediate central nervous system (CNS) and limb polarity. Ectopic expression of Shh in the mouse CNS leads to the activation of floor plate-expressed genes. These results suggest that Shh may play a role in the normal inductive interactions that pattern the ventral CNS.

The splotch-delayed (Spd) mouse mutant carries a point mutation within the paired box of the Pax-3 gene.

The splotch (Sp) mouse mutant displays defects in neural crest cell migration and neural tube closure and serves as a model for the study of spina bifida, exencephaly, and Waardenburg syndrome type I in humans. Recently, we have described alterations in the Pax-3 gene for the radiation-induced Spr and Sp2H alleles and for the original, spontaneously arising Sp allele. Another allele that arose spontaneously at the Sp locus, termed splotch-delayed (Spd), shows a less severe phenotype than the other Sp alleles, including the delayed death of homozygous embryos. To determine the molecular basis underlying this unique phenotype, we have analyzed the integrity of the Pax-3 gene in Spd mutant embryos. Nucleotide sequence analysis of cDNA and genomic clones revealed a G to C transversion at nucleotide 421 of the Pax-3 mRNA transcript, which results in a Gly to Arg substitution at position 42 of the Pax-3 protein (position 9 of the paired domain). The location of the mutated residue, its conservation in all other paired domain-containing proteins described to date, and the nonconservative nature of the substitution suggest that this mutation is responsible for the phenotype observed in the Spd mouse mutant.

A mutation within intron 3 of the Pax-3 gene produces aberrantly spliced mRNA transcripts in the splotch (Sp) mouse mutant.

The splotch (Sp) mouse mutant displays defects in neural tube closure in the form of exencephaly and spina bifida. Recently, mutations in the Pax-3 gene have been described in the radiation-induced Spr and Sp2H alleles. This led us to examine the integrity of the Pax-3 gene and its cellular mRNA transcript in the original, spontaneously arising Sp allele. A complex mutation in the Pax-3 gene including an A-->T transversion at the invariant 3' AG splice acceptor of intron 3 was identified in the Sp/Sp mutant. This genomic mutation abrogates the normal splicing of intron 3, resulting in the generation of four aberrantly spliced mRNA transcripts. Two of these Pax-3 transcripts make use of cryptic 3' splice sites within the downstream exon, generating small deletions which disrupt the reading frame of the transcripts. A third aberrant splicing event results in the deletion of exon 4, while a fourth retains intron 3. These aberrantly spliced mRNA transcripts are not expected to result in functional Pax-3 proteins and are thus responsible for the phenotype observed in the Sp mouse mutant.

Identification and mapping of six microdissected genomic DNA probes to the proximal region of mouse chromosome 1.

Six independent DNA probes, lambda Mm1C-150, lambda Mm1C-153, lambda Mm1C-156, lambda Mm1C-162, lambda Mm1C-163, and lambda Mm1C-165, have been isolated from a library of microdissected fragments from mouse chromosome 1, spanning cytogenetic bands C2 to C5. These DNA probes have been mapped by restriction fragment length polymorphism analysis with respect to 12 marker loci previously assigned to this portion of mouse chromosome 1, in a panel of 251 segregating Mus spretus x C57BL/6J interspecific backcross mice. The gene order and intergene distances were determined by segregation analysis to be centromere- lambda Mm1C-162-11.1 cM-Col3a1-8.8 cM-Len-2-2.6 cM-lambda Mm1C-163-1.6 cM-Fn-1-1.6 cM-Tp-1-0.8 cM-lambda Mm1C-165/Vil-0.4 cM-Inha-2.8 cM-lambda Mm1C-153-2.4 cM-lambda Mm1C-156-1.2 cM-Pax-3-5.6 cM-Akp-3-0.8 cM-Acrg-2.0 cM-Sag-0.5 cM-Col6a3-1.8 cM-lambda Mm1C-150-15.4 cM-Ren1,2. Four of these probes map within a chromosome 1 segment that is homologous to human chromosome 2q. Southern blotting analyses indicate that one of these anonymous probes, lambda Mm1C-165, detects DNA fragments highly conserved across species. These novel polymorphic probes should prove useful for linkage and physical mapping of this chromosomal region.

Splotch (Sp2H), a mutation affecting development of the mouse neural tube, shows a deletion within the paired homeodomain of Pax-3.

The molecular basis of the mouse mutation splotch (Sp), which is associated with spina bifida and exencephaly, was analyzed at three of its alleles, Sp, Sp2H, and Spr. We mapped the paired box gene Pax-3 within the Inha to Akp3 interval, near or at the Sp locus on chromosome 1, and found Pax-3 to be deleted in heterozygous Spr/+ mice. Analysis of genomic DNA and cDNA clones constructed from Sp2H/Sp2H embryos identified a deletion of 32 nucleotides in the Pax-3 mRNA transcript and gene. This deletion maps within the paired homeodomain of PAX-3 and is predicted to create a truncated protein as a result of a newly created termination codon at the deletion breakpoint. Our study provides evidence for a causal link between deletion of the paired homeodomain of Pax-3 and the Sp2H mutation, and infers that Pax-3 plays a key role in normal neural development.

The host resistance locus Bcg is tightly linked to a group of cytoskeleton-associated protein genes that include villin and desmin.

In the mouse, innate resistance or susceptibility to infection with a group of unrelated intracellular parasites which includes, Mycobacteria, Salmonella, and Leishmania is determined by the expression of a single dominant autosomal gene designated Bcg located on the proximal portion of chromosome 1. The gene is expressed at the level of the mature tissue macrophage and influences its capacity to restrict intracellular proliferation of the parasites. We have used restriction fragment length polymorphism analysis in segregating populations of inter- and intraspecific backcross mice and in recombinant inbred strains to position four new marker genes, transition protein 1 (Tp-1), desmin (Des), the alpha subunit of inhibin (Inha), and retinal S-antigen (Sag), in the vicinity of the host resistance locus, Bcg. The gene order for Tp-1, Des, Inha, and Sag was established in an eight-point testcross with respect to anchor loci previously assigned to that portion of mouse chromosome 1 and was found to be centromere-Fn-1-Tp-1-(Vil,Bcg)-Des-Inha-Akp-3-Acrg+ ++-Sag. Two of these new marker genes were found very tightly linked to Bcg: Des was located 0.3 +/- 0.3 cM distal from (Vil,Bcg) and 0.3 +/- 0.3 cM proximal to Inha. Tp-1 mapped 0.8 +/- 0.8 cM proximal and Sag 12.8 +/- 1.7 cM distal to (Vil,Bcg). Tp-1, Des, Inha, and Sag all fall within a large mouse chromosome 1 segment homologous with the telomeric region of the long arm of human chromosome 2 (2q). Our findings indicate that the two closest markers to the host resistance locus, Bcg, encode cytoskeleton-associated proteins which are capable of interaction with actin filaments.

Molecular characterization of a deletion encompassing the splotch mutation on mouse chromosome 1.

We have used a set of markers newly assigned to the proximal portion of mouse chromosome 1 to characterize the chromosomal segment deleted in the splotch-retarded (Spr) mouse mutant. Among nine markers tested in the heterozygote Spr/+mouse, we have identified four genes, Vil, Des, Inha, and Akp-3, which map within the Spr deletion. The closest distal marker to the deletion is the Acrg gene, with the distal deletion breakpoint mapping within the 0.8-cM segment separating Akp-3 and Acrg. The most proximal gene to the Spr deletion is Tp1. The proximal deletion breakpoint maps within the 0.8-cM segment separating Tp1 and Vil. The minimum size of the Spr deletion would therefore be limited to 14 cM, the genetic distance between Vil and Akp-3. The maximum size of the Spr deletion is estimated to be 16 cM, the genetic distance between Tp1 and Acrg.

Genetic control of the survival of murine trisomy 16 fetuses.

A mouse model that allows for the experimental induction of an aneuploid state has been employed to investigate the factors that control the survival of trisomy 16 fetuses. The prevalence of trisomy 16 fetuses on day 15 of gestation was shown to vary significantly with the genetic background of the female parent. The ability to spontaneously abort a trisomy 16 conceptus was shown to be higher in the mouse strain with a low prevalence of trisomy 16, compared to those mouse strains with a high prevalence of trisomy 16. Furthermore, the maternal ability that selects against, or promotes the survival of a trisomic conceptus was shown to be specific for the trisomy in question.

Radioimmunoassays for protein C and factor X. Plasma antigen levels in abnormal hemostatic states.

Specific radioimmunoassays, sensitive to plasma levels of less than 1% of normal, were developed for protein C and Factor X. In 31 normal subjects, mean plasma antigen levels were as follows: protein C, 3.23 +/- 0.79 microgram/mL (2 SD); Factor X, 7.74 +/- 1.81 microgram/mL. In patient son chronic warfarin therapy, protein C and factor X were depressed equivalently: protein C, 42% +/- 20% (of a pooled plasma reference); Factor X, 44% +/- 24%. Protein C antigen fell much more rapidly than Factor X antigen when warfarin therapy was begun, creating an initial period of potential hypercoagulability. In patients with severe liver disease, mean protein C antigen (25% +/- 17%) was lower than Factor X antigen (51% +/- 29%). Protein C antigen levels did not appear to be a sensitive indicator of compensated intravascular coagulation or systemic fibrinolysis induced by infusion of streptokinase. Clinical implications of these findings are discussed.

High-resolution densitometry: analysis of stained albumin bands as a model for electrophoresis of serum proteins.

In discussing the principles of quantitative analysis in thin-layer media, we show that requirements for quantitative analysis are not satisfied when stained protein electrophoretic bands are scanned with a conventional rectangular-slit densitometer. We investigated a high-resolution densitometer based on a linear photodiode array as an alternative analytical tool, using stained electrophoretic bands of radio-labeled human serum albumin as a simplified model for results of serum protein electrophoresis. Identical protein bands scanned with both the high-resolution densitometer and a conventional densitometer were quantified with improved accuracy and precision by the new instrument. We also used the high-resolution densitometer to develop a computer model for performance characteristics of a rectangular-slit densitometer.