Human-Specific NOTCH2NL Genes Affect Notch Signaling and Cortical Neurogenesis.

Genetic changes causing brain size expansion in human evolution have remained elusive. Notch signaling is essential for radial glia stem cell proliferation and is a determinant of neuronal number in the mammalian cortex. We find that three paralogs of human-specific NOTCH2NL are highly expressed in radial glia. Functional analysis reveals that different alleles of NOTCH2NL have varying potencies to enhance Notch signaling by interacting directly with NOTCH receptors. Consistent with a role in Notch signaling, NOTCH2NL ectopic expression delays differentiation of neuronal progenitors, while deletion accelerates differentiation into cortical neurons. Furthermore, NOTCH2NL genes provide the breakpoints in 1q21.1 distal deletion/duplication syndrome, where duplications are associated with macrocephaly and autism and deletions with microcephaly and schizophrenia. Thus, the emergence of human-specific NOTCH2NL genes may have contributed to the rapid evolution of the larger human neocortex, accompanied by loss of genomic stability at the 1q21.1 locus and resulting recurrent neurodevelopmental disorders.

A hidden layer of structural variation in transposable elements reveals potential genetic modifiers in human disease-risk loci.

Genome-wide association studies (GWAS) have been highly informative in discovering disease-associated loci but are not designed to capture all structural variations in the human genome. Using long-read sequencing data, we discovered widespread structural variation within SINE-VNTR-Alu (SVA) elements, a class of great ape-specific transposable elements with gene-regulatory roles, which represents a major source of structural variability in the human population. We highlight the presence of structurally variable SVAs (SV-SVAs) in neurological disease-associated loci, and we further associate SV-SVAs to disease-associated SNPs and differential gene expression using luciferase assays and expression quantitative trait loci data. Finally, we genetically deleted SV-SVAs in the BIN1 and CD2AP Alzheimer's disease-associated risk loci and in the BCKDK Parkinson's disease-associated risk locus and assessed multiple aspects of their gene-regulatory influence in a human neuronal context. Together, this study reveals a novel layer of genetic variation in transposable elements that may contribute to identification of the structural variants that are the actual drivers of disease associations of GWAS loci.

ZNF91 deletion in human embryonic stem cells leads to ectopic activation of SVA retrotransposons and up-regulation of KRAB zinc finger gene clusters.

Transposable element (TE) invasions have shaped vertebrate genomes over the course of evolution. They have contributed an extra layer of species-specific gene regulation by providing novel transcription factor binding sites. In humans, SINE-VNTR-Alu (SVA) elements are one of three still active TE families; approximately 2800 SVA insertions exist in the human genome, half of which are human-specific. TEs are often silenced by KRAB zinc finger (KZNF) proteins recruiting corepressor proteins that establish a repressive chromatin state. A number of KZNFs have been reported to bind SVAs, but their individual contribution to repressing SVAs and their roles in suppressing SVA-mediated gene-regulatory effects remains elusive. We analyzed the genome-wide binding profile for ZNF91 in human cells and found that ZNF91 interacts with the VNTR region of SVAs. Through CRISPR-Cas9-mediated deletion of ZNF91 in human embryonic stem cells, we established that loss of ZNF91 results in increased transcriptional activity of SVAs. In contrast, SVA activation was not observed upon genetic deletion of the ZNF611 gene encoding another strong SVA interactor. Epigenetic profiling confirmed the loss of SVA repression in the absence of ZNF91 and revealed that mainly evolutionary young SVAs gain gene activation-associated epigenetic modifications. Genes close to activated SVAs showed a mild up-regulation, indicating SVAs adopt properties of cis-regulatory elements in the absence of repression. Notably, genome-wide derepression of SVAs elicited the communal up-regulation of KZNFs that reside in KZNF clusters. This phenomenon may provide new insights into the potential mechanisms used by the host genome to sense and counteract TE invasions.

A computational neuroimaging study of reinforcement learning and goal-directed exploration in schizophrenia spectrum disorders.

BACKGROUND: Prior evidence indicates that negative symptom severity and cognitive deficits, in people with schizophrenia (PSZ), relate to measures of reward-seeking and loss-avoidance behavior (implicating the ventral striatum/VS), as well as uncertainty-driven exploration (reliant on rostrolateral prefrontal cortex/rlPFC). While neural correlates of reward-seeking and loss-avoidance have been examined in PSZ, neural correlates of uncertainty-driven exploration have not. Understanding neural correlates of uncertainty-driven exploration is an important next step that could reveal insights to how this mechanism of cognitive and negative symptoms manifest at a neural level. METHODS: We acquired fMRI data from 29 PSZ and 36 controls performing the Temporal Utility Integration decision-making task. Computational analyses estimated parameters corresponding to learning rates for both positive and negative reward prediction errors (RPEs) and the degree to which participates relied on representations of relative uncertainty. Trial-wise estimates of expected value, certainty, and RPEs were generated to model fMRI data. RESULTS: Behaviorally, PSZ demonstrated reduced reward-seeking behavior compared to controls, and negative symptoms were positively correlated with loss-avoidance behavior. This finding of a bias toward loss avoidance learning in PSZ is consistent with previous work. Surprisingly, neither behavioral measures of exploration nor neural correlates of uncertainty in the rlPFC differed significantly between groups. However, we showed that trial-wise estimates of relative uncertainty in the rlPFC distinguished participants who engaged in exploratory behavior from those who did not. rlPFC activation was positively associated with intellectual function. CONCLUSIONS: These results further elucidate the nature of reinforcement learning and decision-making in PSZ and healthy volunteers.

Parity-pair-mixing effects in nonlinear spectroscopy of HDO.

A non-linear spectroscopic study of the HDO molecule is performed in the wavelength range of 1.36-1.42 µm using noise-immune cavity-enhanced optical-heterodyne molecular spectroscopy (NICE-OHMS). More than 100 rovibrational Lamb dips are recorded, with an experimental precision of 2-20 kHz, related to the first overtone of the O-H stretch fundamental of HD16O and HD18O. Significant perturbations, including distortions, shifts, and splittings, have been observed for a number of Lamb dips. These spectral perturbations are traced back to an AC-Stark effect, arising due to the strong laser field applied in all saturation-spectroscopy experiments. The AC-Stark effect mixes parity pairs, that is pairs of rovibrational states whose assignment differs solely in the Kc quantum number, where Kc is part of the standard J K a,K c asymmetric-top rotational label. Parity-pair mixing seems to be especially large for parity pairs with Ka ≥ 3, whereby their energy splittings become as small as a few MHz, resulting in multi-component asymmetric Lamb-dip profiles of gradually increasing complexity. These complex profiles often include crossover resonances. This effect is well known in saturation spectroscopy, but has not been reported in combination with parity-pair mixing. Parity-pair mixing is not seen in H2 16O and H2 18O, because their parity pairs correspond to ortho and para nuclear-spin isomers, whose interaction is prohibited. Despite the frequency shifts observed for HD16O and HD18O, the absolute accuracy of the detected transitions still exceeds that achievable by Doppler-limited techniques.

Molecular Processes in Stress Urinary Incontinence: A Systematic Review of Human and Animal Studies.

Stress urinary incontinence (SUI) is a common and burdensome condition. Because of the large knowledge gap around the molecular processes involved in its pathophysiology, the aim of this review was to provide a systematic overview of genetic variants, gene and protein expression changes related to SUI in human and animal studies. On 5 January 2021, a systematic search was performed in Pubmed, Embase, Web of Science, and the Cochrane library. The screening process and quality assessment were performed in duplicate, using predefined inclusion criteria and different quality assessment tools for human and animal studies respectively. The extracted data were grouped in themes per outcome measure, according to their functions in cellular processes, and synthesized in a narrative review. Finally, 107 studies were included, of which 35 used animal models (rats and mice). Resulting from the most examined processes, the evidence suggests that SUI is associated with altered extracellular matrix metabolism, estrogen receptors, oxidative stress, apoptosis, inflammation, neurodegenerative processes, and muscle cell differentiation and contractility. Due to heterogeneity in the studies (e.g., in examined tissues), the precise contribution of the associated genes and proteins in relation to SUI pathophysiology remained unclear. Future research should focus on possible contributors to these alterations.

Evolution of Human Brain Size-Associated NOTCH2NL Genes Proceeds toward Reduced Protein Levels.

Ever since the availability of genomes from Neanderthals, Denisovans, and ancient humans, the field of evolutionary genomics has been searching for protein-coding variants that may hold clues to how our species evolved over the last ∼600,000 years. In this study, we identify such variants in the human-specific NOTCH2NL gene family, which were recently identified as possible contributors to the evolutionary expansion of the human brain. We find evidence for the existence of unique protein-coding NOTCH2NL variants in Neanderthals and Denisovans which could affect their ability to activate Notch signaling. Furthermore, in the Neanderthal and Denisovan genomes, we find unusual NOTCH2NL configurations, not found in any of the modern human genomes analyzed. Finally, genetic analysis of archaic and modern humans reveals ongoing adaptive evolution of modern human NOTCH2NL genes, identifying three structural variants acting complementary to drive our genome to produce a lower dosage of NOTCH2NL protein. Because copy-number variations of the 1q21.1 locus, encompassing NOTCH2NL genes, are associated with severe neurological disorders, this seemingly contradicting drive toward low levels of NOTCH2NL protein indicates that the optimal dosage of NOTCH2NL may have not yet been settled in the human population.

Hyperfine-Resolved Near-Infrared Spectra of H217O.

Huge efforts have recently been taken in the derivation of accurate compilations of rovibrational energies of water, one of the most important reference systems in spectroscopy. Such precision is desirable for all water isotopologues, although their investigation is challenged by hyperfine effects in their spectra. Frequency-comb locked noise-immune cavity-enhanced optical-heterodyne molecular spectroscopy (NICE-OHMS) allows for achieving high sensitivity, resolution, and accuracy. This technique has been employed to resolve the subtle hyperfine splittings of rovibrational transitions of H217O in the near-infrared region. Simulation and interpretation of the H217O saturation spectra have been supported by coupled-cluster calculations performed with large basis sets and accounting for high-level corrections. Experimental 17O hyperfine parameters are found in excellent agreement with the corresponding computed values. The need of including small hyperfine effects in the analysis of H217O spectra has been demonstrated together with the ability of the computational strategy employed for providing quantitative predictions of the corresponding parameters.

An evolutionary arms race between KRAB zinc-finger genes ZNF91/93 and SVA/L1 retrotransposons.

Throughout evolution primate genomes have been modified by waves of retrotransposon insertions. For each wave, the host eventually finds a way to repress retrotransposon transcription and prevent further insertions. In mouse embryonic stem cells, transcriptional silencing of retrotransposons requires KAP1 (also known as TRIM28) and its repressive complex, which can be recruited to target sites by KRAB zinc-finger (KZNF) proteins such as murine-specific ZFP809 which binds to integrated murine leukaemia virus DNA elements and recruits KAP1 to repress them. KZNF genes are one of the fastest growing gene families in primates and this expansion is hypothesized to enable primates to respond to newly emerged retrotransposons. However, the identity of KZNF genes battling retrotransposons currently active in the human genome, such as SINE-VNTR-Alu (SVA) and long interspersed nuclear element 1 (L1), is unknown. Here we show that two primate-specific KZNF genes rapidly evolved to repress these two distinct retrotransposon families shortly after they began to spread in our ancestral genome. ZNF91 underwent a series of structural changes 8-12 million years ago that enabled it to repress SVA elements. ZNF93 evolved earlier to repress the primate L1 lineage until ∼12.5 million years ago when the L1PA3-subfamily of retrotransposons escaped ZNF93's restriction through the removal of the ZNF93-binding site. Our data support a model where KZNF gene expansion limits the activity of newly emerged retrotransposon classes, and this is followed by mutations in these retrotransposons to evade repression, a cycle of events that could explain the rapid expansion of lineage-specific KZNF genes.

Immediate early genes in social insects: a tool to identify brain regions involved in complex behaviors and molecular processes underlying neuroplasticity.

Social insects show complex behaviors and master cognitive tasks. The underlying neuronal mechanisms, however, are in most cases only poorly understood due to challenges in monitoring brain activity in freely moving animals. Immediate early genes (IEGs) that get rapidly and transiently expressed following neuronal stimulation provide a powerful tool for detecting behavior-related neuronal activity in vertebrates. In social insects, like honey bees, and in insects in general, this approach is not yet routinely established, even though these genes are highly conserved. First studies revealed a vast potential of using IEGs as neuronal activity markers to analyze the localization, function, and plasticity of neuronal circuits underlying complex social behaviors. We summarize the current knowledge on IEGs in social insects and provide ideas for future research directions.

Motivational deficits in schizophrenia relate to abnormalities in cortical learning rate signals.

Individuals from across the psychosis spectrum display impairments in reinforcement learning. In some individuals, these deficits may result from aberrations in reward prediction error (RPE) signaling, conveyed by dopaminergic projections to the ventral striatum (VS). However, there is mounting evidence that VS RPE signals are relatively intact in medicated people with schizophrenia (PSZ). We hypothesized that, in PSZ, reinforcement learning deficits often are not related to RPE signaling per se but rather their impact on learning and behavior (i.e., learning rate modulation), due to dysfunction in anterior cingulate and dorsomedial prefrontal cortex (dmPFC). Twenty-six PSZ and 23 healthy volunteers completed a probabilistic reinforcement learning paradigm with occasional, sudden, shifts in contingencies. Using computational modeling, we found evidence of an impairment in trial-wise learning rate modulation (α) in PSZ before and after a reinforcement contingency shift, expressed most in PSZ with more severe motivational deficits. In a subsample of 22 PSZ and 22 healthy volunteers, we found little evidence for between-group differences in VS RPE and dmPFC learning rate signals, as measured with fMRI. However, a follow-up psychophysiological interaction analysis revealed decreased dmPFC-VS connectivity concurrent with learning rate modulation, most prominently in individuals with the most severe motivational deficits. These findings point to an impairment in learning rate modulation in PSZ, leading to a reduced ability to adjust task behavior in response to unexpected outcomes. At the level of the brain, learning rate modulation deficits may be associated with decreased involvement of the dmPFC within a greater RL network.

A New Implanted Posterior Tibial Nerve Stimulator for the Treatment of Overactive Bladder Syndrome: 3-Month Results of a Novel Therapy at a Single Center.

PURPOSE: This study was designed to investigate the safety and performance of a new implantable system for tibial nerve stimulation for overactive bladder symptoms. MATERIALS AND METHODS: A battery-free stimulation device for tibial nerve stimulation (BlueWind Medical, Herzliya, Israel) was implanted in 15 patients. Safety and efficacy assessments were done at 3 months after activation with a 3-day bladder diary, a 24-hour pad test and 2 quality of life questionnaires. RESULTS: Two males and 13 females were enrolled in the study. Mean age was 54 years (range 19 to 72). Five of 15 patients were previously treated with percutaneous tibial nerve stimulation and 12 experienced urgency urinary incontinence. Median operative time was 34 minutes. At 3 months of followup a significant change was seen in 24-hour frequency from a mean ± SD of 11.8 ± 3.5 to 8.1 ± 2.0 times per day (p = 0.002), the number of severe urinary urgency episodes from 6.5 ± 5.1 to 2.0 ± 2.1 times per day (p = 0.002), the number of severe incontinence episodes from 2.8 ± 5.2 to 0.3 ± 0.4 episodes per day (p = 0.017), urinary loss per day from 243 ± 388 to 39 ± 55 gm (p = 0.038) and improvement in quality of life. After implantation, 3 patients received prolonged antibiotic treatment and 3 received pain medication for 1 week. In 1 patient the device was explanted due to pain and swelling suspicious for infection, although tissue cultures did not reveal a bacterial infection. CONCLUSIONS: This novel posterior tibial nerve stimulator is safe and easy to implant with good clinical results.

Constraints on Oscillation Parameters from ν_{e} Appearance and ν_{µ} Disappearance in NOvA.

Results are reported from an improved measurement of ν_{µ}→ν_{e} transitions by the NOvA experiment. Using an exposure equivalent to 6.05×10^{20} protons on target, 33 ν_{e} candidates are observed with a background of 8.2±0.8 (syst.). Combined with the latest NOvA ν_{µ} disappearance data and external constraints from reactor experiments on sin^{2}2θ_{13}, the hypothesis of inverted mass hierarchy with θ_{23} in the lower octant is disfavored at greater than 93% C.L. for all values of δ_{CP}.

Retinoic acid-dependent and -independent gene-regulatory pathways of Pitx3 in meso-diencephalic dopaminergic neurons.

Development of meso-diencephalic dopamine (mdDA) neurons requires the combined actions of the orphan nuclear receptor Nurr1 and the paired-like homeobox transcription factor Pitx3. Whereas all mdDA neurons require Nurr1 for expression of Th and survival, dependence on Pitx3 is displayed only by the mdDA subpopulation that will form the substantia nigra (SNc). Previously, we have demonstrated that Pitx3(-/-) embryos lack the expression of the retinoic acid (RA)-generating enzyme Ahd2, which is normally selectively expressed in the Pitx3-dependent DA neurons of the SNc. Restoring RA signaling in Pitx3(-/-) embryos revealed a selective dependence of SNc neurons on the presence of RA for differentiation into Th-positive neurons and maintenance throughout embryonic development. Whereas these data are suggestive of an important developmental role for RA in neurons of the SNc, it remained unclear whether other Nurr1 and Pitx3 target genes depend on RA signaling in a manner similar to Th. In the search for genes that were affected in Pitx3-deficient mdDA neurons and restored upon embryonic RA treatment, we provide evidence that Delta-like 1, D2R (Drd2) and Th are regulated by Pitx3 and RA signaling, which influences the mdDA terminal differentiated phenotype. Furthermore, we show that regulation of Ahd2-mediated RA signaling represents only one aspect of the Pitx3 downstream cascade, as Vmat2, Dat, Ahd2 (Aldh1a1), En1, En2 and Cck were unaffected by RA treatment and are (subset) specifically modulated by Pitx3. In conclusion, our data reveal several RA-dependent and -independent aspects of the Pitx3-regulated gene cascade, suggesting that Pitx3 acts on multiple levels in the molecular subset-specification of mdDA neurons.

Emergence of the ZNF675 Gene During Primate Evolution-Influenced Human Neurodevelopment Through Changing HES1 Autoregulation.

In this study, we investigated recurrent copy number variations (CNVs) in the 19p12 locus, which are associated with neurodevelopmental disorders. The two genes in this locus, ZNF675 and ZNF681, arose via gene duplication in primates, and their presence in several pathological CNVs in the human population suggests that either or both of these genes are required for normal human brain development. ZNF675 and ZNF681 are members of the Krüppel-associated box zinc finger (KZNF) protein family, a class of transcriptional repressors important for epigenetic silencing of specific genomic regions. About 170 primate-specific KZNFs are present in the human genome. Although KZNFs are primarily associated with repressing retrotransposon-derived DNA, evidence is emerging that they can be co-opted for other gene regulatory processes. We show that genetic deletion of ZNF675 causes developmental defects in cortical organoids, and our data suggest that part of the observed neurodevelopmental phenotype is mediated by a gene regulatory role of ZNF675 on the promoter of the neurodevelopmental gene Hes family BHLH transcription factor 1 (HES1). We also find evidence for the recently evolved regulation of genes involved in neurological disorders, microcephalin 1 and sestrin 3. We show that ZNF675 interferes with HES1 auto-inhibition, a process essential for the maintenance of neural progenitors. As a striking example of how some KZNFs have integrated into preexisting gene expression networks, these findings suggest the emergence of ZNF675 has caused a change in the balance of HES1 autoregulation. The association of ZNF675 CNV with human developmental disorders and ZNF675-mediated regulation of neurodevelopmental genes suggests that it evolved into an important factor for human brain development.

Six Steps towards a Spatial Design for Large-Scale Pollinator Surveillance Monitoring.

Despite the importance of pollinators to ecosystem functioning and human food production, comprehensive pollinator monitoring data are still lacking across most regions of the world. Policy-makers have recently prioritised the development of large-scale monitoring programmes for pollinators to better understand how populations respond to land use, environmental change and restoration measures in the long term. Designing such a monitoring programme is challenging, partly because it requires both ecological knowledge and advanced knowledge in sampling design. This study aims to develop a conceptual framework to facilitate the spatial sampling design of large-scale surveillance monitoring. The system is designed to detect changes in pollinator species abundances and richness, focusing on temperate agroecosystems. The sampling design needs to be scientifically robust to address questions of agri-environmental policy at the scales of interest. To this end, we followed a six-step procedure as follows: (1) defining the spatial sampling units, (2) defining and delimiting the monitoring area, (3) deciding on the general sampling strategy, (4) determining the sample size, (5) specifying the sampling units per sampling interval, and (6) specifying the pollinator survey plots within each sampling unit. As a case study, we apply this framework to the "Wild bee monitoring in agricultural landscapes of Germany" programme. We suggest this six-step procedure as a conceptual guideline for the spatial sampling design of future large-scale pollinator monitoring initiatives.

All paths lead to hubs in the spectroscopic networks of water isotopologues H216O and H218O.

Network theory has fundamentally transformed our comprehension of complex systems, catalyzing significant advances across various domains of science and technology. In spectroscopic networks, hubs are the quantum states involved in the largest number of transitions. Here, utilizing network paths probed via precision metrology, absolute energies have been deduced, with at least 10-digit accuracy, for almost 200 hubs in the experimental spectroscopic networks of H216O and H218O. These hubs, lying on the ground vibrational states of both species and the bending fundamental of H216O, are involved in tens of thousands of observed transitions. Relying on the same hubs and other states, benchmark-quality line lists have been assembled, which supersede and improve, by three orders of magnitude, the accuracy of the massive amount of data reported in hundreds of papers dealing with Doppler-limited spectroscopy. Due to the omnipresence of water, these ultraprecise line lists could be applied to calibrate high-resolution spectra and serve ongoing and upcoming space missions.

Brain exposure to SARS-CoV-2 virions perturbs synaptic homeostasis.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is associated with short- and long-term neurological complications. The variety of symptoms makes it difficult to unravel molecular mechanisms underlying neurological sequalae after coronavirus disease 2019 (COVID-19). Here we show that SARS-CoV-2 triggers the up-regulation of synaptic components and perturbs local electrical field potential. Using cerebral organoids, organotypic culture of human brain explants from individuals without COVID-19 and post-mortem brain samples from individuals with COVID-19, we find that neural cells are permissive to SARS-CoV-2 to a low extent. SARS-CoV-2 induces aberrant presynaptic morphology and increases expression of the synaptic components Bassoon, latrophilin-3 (LPHN3) and fibronectin leucine-rich transmembrane protein-3 (FLRT3). Furthermore, we find that LPHN3-agonist treatment with Stachel partially restored organoid electrical activity and reverted SARS-CoV-2-induced aberrant presynaptic morphology. Finally, we observe accumulation of relatively static virions at LPHN3-FLRT3 synapses, suggesting that local hindrance can contribute to synaptic perturbations. Together, our study provides molecular insights into SARS-CoV-2-brain interactions, which may contribute to COVID-19-related neurological disorders.

Long-term testicular volume after orchiopexy at diagnosis of acquired undescended testis.

PURPOSE: We studied long-term outcomes of orchiopexy at diagnosis of acquired undescended testes using ultrasound to determine testicular volume. MATERIALS AND METHODS: Patients who had undergone orchiopexy for acquired undescended testis at diagnosis were recruited to assess testicular volume. Testis volume was measured by ultrasound and compared with recently developed normative values for testicular size. For young adults (older than 18 years) volumes were grouped and compared to normative values reported in the literature. In all unilateral cases testicular volume was compared with its counterpart. RESULTS: A total of 155 patients 5.1 to 26.6 years old (181 acquired undescended testes) were included in the study. Mean ± SD followup was 6.6 ± 3.8 years (range 1.4 to 15.5). For all patients 18 years old or younger (125 patients, 143 testes) operated testis volume was 0.1 to 12.7 ml (mean ± SD 2.5 ± 2.9), which was significantly smaller than the normative values (50th percentile) for the same age (p <0.001). Mean ± SD testis volume in young adults (38 testes) was 8.1 ± 3.7 ml, compared to a mean volume of 13.4 ml reported in the literature (p <0.001). In unilateral cases the mean volume of the testes fixed by orchiopexy differed significantly from their counterparts (3.4 ± 3.3 ml vs 4.6 ± 4.6 ml, p <0.001). CONCLUSIONS: The long-term volumes at diagnosis of acquired undescended testes after orchiopexy were significantly less than the normative values at all ages. In unilateral cases the volumes were also significantly less compared to the contralateral testes.

Increased reactivity and in vitro cell response of titanium based implant surfaces after anodic oxidation.

In the quest for improved bone growth and attachment around dental implants, chemical surface modifications are one possibility for future developments. The biological properties of titanium based materials can be further enhanced with methods like anodic polarization to produce an active rather than a passive titanium oxide surface. Here we investigate the formation of hydroxide groups on sand blasted and acid etched titanium and titanium-zirconium alloy surfaces after anodic polarization in an alkaline solution. X-ray photoelectron spectroscopy shows that the activated surfaces had increased reactivity. Furthermore the activated surfaces show up to threefold increase in OH(-) concentration in comparison to the original surface. The surface parameters Sa, Sku, Sdr and Ssk were more closely correlated to time and current density for titanium than for titanium-zirconium. Studies with MC3T3-E1 osteoblastic cells showed that OH(-) activated surfaces increased mRNA levels of osteocalcin and collagen-I.