The Effect of Helium Ion Radiation on the Material Properties of Bone.

Ionizing radiation, from both space and radiation therapy, is known to affect bone health. While there have been studies investigating changes in bone density and microstructure from radiation exposure, the effects of radiation on material properties are unknown. The current study addresses this gap by assessing bone material property changes in rats exposed to helium-4 radiation through spherical micro-indentation. Rats were exposed to a single dose of 0, 5, and 25 cGy whole body helium-4 radiation. Animals were euthanized at 7, 30, 90, or 180-days after exposure. Spherical micro-indentation was performed on axial cross sections of the femur cortical bone to determine instantaneous and relaxed shear moduli. At 90-days after exposure, the 25 cGy exposure caused a significant decline in shear modulus compared to control and 5 cGy groups. The instantaneous modulus decreased 33% and the relaxed modulus decreased 32% as compared to the sham group. This decline was followed by a recovery of both moduli, which was observed by 180-days after exposure; at 180 days, the moduli were no longer statistically different from those at 7 or 30 days. The observed decrease at 90 days, followed by recovery to baseline levels, can be attributed to the biological mechanisms involved in bone formation that were affected by radiation, bone turnover, and systemic changes in hormones due to radiation exposure. Continued assessment of the mechanisms that drive such a response in material properties may enable identification of pathways for therapeutic countermeasures against radiation exposure.

Bone material strength in normoglycemic and hyperglycemic black and white older adults.

This cross-sectional study assessed cortical bone properties via impact microindentation in adults with normoglycemia, prediabetes, and early-stage T2D. Bone material strength index was stable across the glycemia categories in whites but it declined in blacks. Blacks may be more susceptible than whites to impaired cortical bone properties in early diabetes. INTRODUCTION: Individuals with long-standing type 2 diabetes (T2D) have altered cortical bone material properties as determined by impact microindentation. This cross-sectional study was done to determine whether altered cortical bone material properties could be detected in adults with prediabetes or early-stage T2D. METHODS: Men and postmenopausal women aged ≥ 50 years with no diabetes (50 white, 6 black), prediabetes (75 white, 13 black), and T2D of ≤ 5 years duration (24 white and 16 black) had assessments of bone material strength index (BMSi) by impact microindentation, trabecular bone score (TBS), and bone mineral density (BMD) by DXA and the advanced glycation end product, urine pentosidine. RESULTS: The association between glycemia category and BMSi differed by race (interaction p = 0.037). In the whites, BMSi did not differ across the glycemia categories, after adjustment for age, sex, and BMI (no diabetes 76.3 ± 1.6 (SEM), prediabetes 77.2 ± 1.3, T2D 76.2 ± 2.5, ANCOVA p = 0.887). In contrast, in the blacks, BMSi differed (ANCOVA p = 0.020) and was significantly lower in subjects with T2D than in those with prediabetes (p < 0.05) and no diabetes (p < 0.05) (mean ± SEM BMSi in no diabetes 86.0 ± 4.3, prediabetes 91.0 ± 3.2, and T2D 71.6 ± 2.9). Neither TBS nor urine pentosidine differed significantly across the glycemia categories in either whites or blacks. CONCLUSIONS: These findings suggest different associations of glycemia with cortical bone material properties in blacks and whites, with blacks possibly being more susceptible to impaired cortical bone properties than whites in early diabetes. A larger study is needed to verify these observations.

Structure, composition, mechanics and growth of spines of the dorsal fin of blue tilapia Oreochromis aureus and common carp Cyprinus carpio.

The structural, compositional and mechanical properties of the spines of the dorsal fin in mature anosteocytic blue tilapia Oreochromis aureus and osteocytic common carp Cyprinus carpio are described, as well as their temporal growth pattern and regenerative capacities. The three-dimensional architecture of both spines, from macro to sub-micron levels, is shown to be axially oriented and therefore highly anisotropic and the spines of both species are able to regenerate after partial amputation.

Measuring the mechanical properties of plant cells by combining micro-indentation with osmotic treatments.

Growth in plants results from the interaction between genetic and signalling networks and the mechanical properties of cells and tissues. There has been a recent resurgence in research directed at understanding the mechanical aspects of growth, and their feedback on genetic regulation. This has been driven in part by the development of new micro-indentation techniques to measure the mechanical properties of plant cells in vivo. However, the interpretation of indentation experiments remains a challenge, since the force measures results from a combination of turgor pressure, cell wall stiffness, and cell and indenter geometry. In order to interpret the measurements, an accurate mechanical model of the experiment is required. Here, we used a plant cell system with a simple geometry, Nicotiana tabacum Bright Yellow-2 (BY-2) cells, to examine the sensitivity of micro-indentation to a variety of mechanical and experimental parameters. Using a finite-element mechanical model, we found that, for indentations of a few microns on turgid cells, the measurements were mostly sensitive to turgor pressure and the radius of the cell, and not to the exact indenter shape or elastic properties of the cell wall. By complementing indentation experiments with osmotic experiments to measure the elastic strain in turgid cells, we could fit the model to both turgor pressure and cell wall elasticity. This allowed us to interpret apparent stiffness values in terms of meaningful physical parameters that are relevant for morphogenesis.

A Traumatic Impact Immediately Changes the Mechanical Properties of Articular Cartilage.

OBJECTIVE: To investigate whether and how a single traumatic impact changes the mechanical properties of talar articular cartilage. DESIGN: A marble was placed on the joint surface and a weight was dropped on both medial and lateral caprine talus to create a well-defined single focal impact. The mechanical properties of intact and impacted talar cartilage were measured with a micro-indenter. Elastic (storage) and viscous (loss) moduli were determined by oscillatory ramp and dynamic mechanical analysis protocols. RESULTS: We found significant differences between ankles and within the same ankle joint, with the medial talus having significantly higher storage- and loss moduli than the lateral talus. The storage- and loss moduli of intact articular cartilage increased with greater indentation depths. However, postimpact the storage- and loss moduli were significantly and consistently lower in all specimens indicating immediate posttraumatic damage. The deeper regions of talar cartilage were less affected by the impact than the more superficial regions. CONCLUSIONS: A single traumatic impact results in an immediate and significant decrease of storage- and loss moduli. Further research must focus on the development of non- or minimally invasive diagnostic tools to address the exact microdamage caused by the impact. We speculate that the traumatic impact damaged the collagen fibers that confine the water-binding proteoglycans and thereby decreasing the hydrostatic pressure of cartilage. As part of the treatment directly after a trauma, one could imagine a reduction or restriction of peak loads to prevent the progression of the cascade towards PTOA of the ankle joint.

Microhardness and Microstructure Analysis of the LPBF Additively Manufactured 18Ni300.

This research focuses on analysing the 18Ni300 maraging steel produced through laser powder bed fusion. Specifically, it aims to examine the phase components using X-ray diffraction, the microstructure through scanning electron microscopy, and the hardness of the different structures present in the manufactured material. The primary goal is to meticulously analyse the material and its microstructures. By doing so, a correlation between the hardness and each structure type, be it cellular or columnar, can be established. This will allow us to pinpoint any defects in the material before any surface chemical treatment is carried out and facilitate a thorough examination of its microstructure. A consistent pattern emerges across the samples through systematic measurement of microhardness distribution in various locations and detailed examination of the structure. The findings of the study reveal that the hardness of cellular and columnar structures exhibits a significant variation based on the location of the measurement about cell boundaries. The hardness value is notably higher in the combination of cellular and multiple layers, as the data indicate.

Characterization of the Vertical Stiffness Gradient in Cadaveric Human and Excised Canine Larynges.

The elastic properties of the folds govern the characteristics of vocal fold vibrations. This study addresses existing gaps by investigating the Young's modulus along the anterior-posterior direction in excised canine and cadaveric human vocal folds. Micro-indentation testing was conducted on six excised canines and three cadaveric human larynges. Multiple points along the medial glottal wall were indented to determine force-displacement, stress-strain relationships, and Young's modulus as a function of Green's strain. A vertical stiffness gradient was consistently observed in both canine and human samples, with higher stiffness in the inferior aspect compared with the superior aspect. The stiffness increased toward both the anterior and posterior directions from the mid-coronal plane, with a more pronounced increase at the posterior edge. Human vocal folds generally exhibited lower stiffness at low strains but were comparable to canine vocal folds at higher strains. These findings suggest that the canine larynx model is a reasonable representation of the human laryngeal tissues' elastic property trends. This analysis of the vertical stiffness gradient in canine and human vocal folds provides valuable data for improving experimental and numerical models of phonation.

Crosslinking of Bovine Gelatin Gels by Genipin Revisited Using Ferrule-Top Micro-Indentation.

(1) Background: Gelatin is widely used in food science, bioengineering, and as a sealant. However, for most of those applications, the mechanical properties of gelatin gels need to be improved by means of physical or chemical crosslinking. Among the used chemical agents, genipin allows low cytotoxicity in addition to improved Young's modulus. However, the mechanical properties of gelatin-genipin gels have only been investigated at the macroscale, and there is no knowledge of the influence of the genipin concentration on the surface homogeneity of Young's modulus. (2) Methods: To this aim, the influence of genipin concentration on Young's modulus of gelatin gels was investigated by means of ferrule-top micro-indentation. The data were compared with storage moduli obtained by shear rheology data. (3) Results: Ferrule-top indentation measurements allowed us to show that Young's moduli of gelatin-genipin gels increase up to a plateau value after approximately 12 mg/mL in genipin and 4 h of crosslinking. Young's moduli distribute with high homogeneity over 80 µm × 80 µm surface areas and are consistent with the storage moduli obtained by shear rheology. (4) Conclusions: It has been shown that ferrule-top indentation data fitted with the Hertz model yield Young's moduli of gelatin-genipin gels which are consistent with the storage moduli obtained by characterization at the macroscale using shear rheometry. In addition, Young's moduli are homogenously distributed (with some irregularities at the highest genipin concentrations) and can be increased by two orders of magnitude with respect to the uncrosslinked gel.

Load-induced fluid pressurisation in hydrogel systems before and after reinforcement by melt-electrowritten fibrous meshes.

Fluid pressure develops transiently within mechanically-loaded, cell-embedding hydrogels, but its magnitude depends on the intrinsic material properties of the hydrogel and cannot be easily altered. The recently developed melt-electrowriting (MEW) technique enables three-dimensional printing of structured fibrous mesh with small fibre diameter (20 µm). The MEW mesh with 20 µm fibre diameter can synergistically increase the instantaneous mechanical stiffness of soft hydrogels. However, the reinforcing mechanism of the MEW meshes is not well understood, and may involve load-induced fluid pressurisation. Here, we examined the reinforcing effect of MEW meshes in three hydrogels: gelatin methacryloyl (GelMA), agarose and alginate, and the role of load-induced fluid pressurisation in the MEW reinforcement. We tested the hydrogels with and without MEW mesh (i.e., hydrogel alone, and MEW-hydrogel composite) using micro-indentation and unconfined compression, and analysed the mechanical data using biphasic Hertz and mixture models. We found that the MEW mesh altered the tension-to-compression modulus ratio differently for hydrogels that are cross-linked differently, which led to a variable change to their load-induced fluid pressurisation. MEW meshes only enhanced the fluid pressurisation for GelMA, but not for agarose or alginate. We speculate that only covalently cross-linked hydrogels (GelMA) can effectively tense the MEW meshes, thereby enhancing the fluid pressure developed during compressive loading. In conclusion, load-induced fluid pressurisation in selected hydrogels was enhanced by MEW fibrous mesh, and may be controlled by MEW mesh of different designs in the future, thereby making fluid pressure a tunable cell growth stimulus for tissue engineering involving mechanical stimulation.

Evaluation of Cross-Linked Polyamide 6 Micro-Indentation Properties: TAIC Concentration and Electron Radiation Intensity.

Nowadays, technical practice puts emphasis on improving selected material properties of polymers which could lead to new applications. Material properties can be modified in numerous ways, among which is radiation treatment. This study looks into the influence of beta radiation on several properties of polyamide 6, e.g., indentation hardness, modulus and creep. Main changeable parameters were the concentration of triallyl isocyanurate (TAIC), which promotes cross-linking, and intensity of radiation. The concentration was in the range from 2 to 6 wt.%, while the radiation dose was 0, 66, 99 and 132 kGy. The treated materials were measured for indentation hardness, modulus and creep. Degree of cross-linking was verified by thermo-mechanical analysis (TMA), while degradation processes was investigated by Fourier-transform infrared spectroscopy (FTIR). The results indicate that electron radiation positively affects the tested material properties. The best results were seen in polyamide with 6 wt.% of TAIC, which demonstrated a 38% improvement in mechanical properties after exposure to 132 kGy. This improvement in properties affects the final parts and their application (e.g., in the automotive industry-engine parts; in electrical engineering-insulation of wires and cables; and in industry-pipes for underfloor heating, etc.).

Multi-length scale strengthening and cytocompatibility of ultra high molecular weight polyethylene bio-composites by functionalized carbon nanotube and hydroxyapatite reinforcement.

The mechanical properties, such as hardness and elastic modulus, of ultra-high molecular weight polyethylene (UHMWPE) composites for acetabular cup liner are improved by adding hydroxyapatite (HAp) and carbon nanotubes (CNT). However, the weak adhesion of HAp (H) and CNT (C) with UHMWPE (U) limits the enhancement of mechanical properties. Thus, the surface of these reinforcements is silane-treated to improve the adhesion with polymer via Si-O and C=O bonds, as evidenced from spectroscopy techniques. An increased dispersion and interfacial adhesion of functionalized HAp (fH) and CNT (fC) with the polymer matrix is confirmed by nearly two-fold increased reinforcement fraction (Rf: 0.55) of U-10 wt% fHAp-2 wt.% fCNT (U10fH2fC) in comparison to U-10 wt% HAp-2 wt.% CNT (U10H2C). Additionally, Voronoi Tessellation (VT) on SEM micrographs of U10H2C and U10fH2fC revealed the dispersion of functionalized CNTs in U10fH2fC with a center-to-center distance of 0.076 µm, which is 74% higher for unfunctionalized CNT in U10H2C. The multilength scale strengthening of the UHMWPE matrix is confirmed from atomic level modification via functionalization of fillers which effectively adhered to the polymer chain on a micro-scale level. A uniform distribution of CNTs rendered increased crystallinity (+28%) of U10fH2fC, which in turn resulted in significant improvement in bulk mechanical properties (18%, 49%, and 12% increased hardness (148.1 MPa), elastic modulus (3.51 GPa) and tensile elastic modulus (219.8 MPa), respectively) in comparison to that of U10H2C. Functionalized-HAp/CNT reinforced UHMWPE composites maintained its cytocompatibility in the MTT test and fluorescence microscopy, affirming their potential employment as acetabular cup liners for hip joint arthroplasty.

Compositional Effects on Indentation Mechanical Properties of Chemically Strengthened TiO2-Doped Soda Lime Silicate Glasses.

TiO2 is an important oxide for property modifications in the conventional soda lime silicate glass family. It offers interesting optical and mechanical properties, for instance, by substituting heavy metals such as lead in consumer glasses. The compositional effects on the hardness, reduced elastic modulus and crack resistance as determined by indentation of chemically strengthened (CS) TiO2-doped soda lime silicate glass was studied in the current paper. The CS, which was performed by a K+ for Na+ ion exchange in a molten KNO3 salt bath at 450 °C for 15 h, yielded significant changes in the indentation mechanical properties. The hardness of the glass samples increased, and this was notably dependent on the SiO2, CaO and TiO2 content. The reduced elastic modulus was less affected by the CS but showed decrease for most samples. The crack resistance, an important property in many applications where glasses are subjected to contact damage, showed very different behaviors among the series. Only one of the series did significantly improve the crack resistance where low CaO content, high TiO2 content, high molar volume and increased elastic deformation favored an increased crack resistance.

Cell wall and cytoskeletal contributions in single cell biomechanics of Nicotiana tabacum.

Studies on the mechanics of plant cells usually focus on understanding the effects of turgor pressure and properties of the cell wall (CW). While the functional roles of the underlying cytoskeleton have been studied, the extent to which it contributes to the mechanical properties of cells is not elucidated. Here, we study the contributions of the CW, microtubules (MTs) and actin filaments (AFs), in the mechanical properties of Nicotiana tabacum cells. We use a multiscale biomechanical assay comprised of atomic force microscopy and micro-indentation in solutions that (i) remove MTs and AFs and (ii) alter osmotic pressures in the cells. To compare measurements obtained by the two mechanical tests, we develop two generative statistical models to describe the cell's behaviour using one or both datasets. Our results illustrate that MTs and AFs contribute significantly to cell stiffness and dissipated energy, while confirming the dominant role of turgor pressure.

Influence of Femtosecond Laser Modification on Biomechanical and Biofunctional Behavior of Porous Titanium Substrates.

Bone resorption and inadequate osseointegration are considered the main problems of titanium implants. In this investigation, the texture and surface roughness of porous titanium samples obtained by the space holder technique were modified with a femtosecond Yb-doped fiber laser. Different percentages of porosity (30, 40, 50, and 60 vol.%) and particle range size (100-200 and 355-500 µm) were compared with fully-dense samples obtained by conventional powder metallurgy. After femtosecond laser treatment the formation of a rough surface with micro-columns and micro-holes occurred for all the studied substrates. The surface was covered by ripples over the micro-metric structures. This work evaluates both the influence of the macro-pores inherent to the spacer particles, as well as the micro-columns and the texture generated with the laser, on the wettability of the surface, the cell behavior (adhesion and proliferation of osteoblasts), micro-hardness (instrumented micro-indentation test, P-h curves) and scratch resistance. The titanium sample with 30 vol.% and a pore range size of 100-200 µm was the best candidate for the replacement of small damaged cortical bone tissues, based on its better biomechanical (stiffness and yield strength) and biofunctional balance (bone in-growth and in vitro osseointegration).

Have female twisted-wing parasites (Insecta: Strepsiptera) evolved tolerance traits as response to traumatic penetration?

Traumatic insemination describes an unusual form of mating during which a male penetrates the body wall of its female partner to inject sperm. Females unable to prevent traumatic insemination have been predicted to develop either traits of tolerance or of resistance, both reducing the fitness costs associated with the male-inflicted injury. The evolution of tolerance traits has previously been suggested for the bed bug. Here we present data suggesting that tolerance traits also evolved in females of the twisted-wing parasite species Stylops ovinae and Xenos vesparum. Using micro-indentation experiments and confocal laser scanning microscopy, we found that females of both investigated species possess a uniform resilin-rich integument that is notably thicker at penetration sites than at control sites. As the thickened cuticle does not seem to hamper penetration by males, we hypothesise that thickening of the cuticle resulted in reduced penetration damage and loss of haemolymph and in improved wound sealing. To evaluate the evolutionary relevance of the Stylops-specific paragenital organ and penis shape variation in the context of inter- and intraspecific competition, we conducted attraction and interspecific mating experiments, as well as a geometric-morphometric analysis of S. ovinae and X. vesparum penises. We found that S. ovinae females indeed attract sympatrically distributed congeneric males. However, only conspecific males were able to mate. In contrast, we did not observe any heterospecific male attraction by Xenos females. We therefore hypothesise that the paragenital organ in the genus Stylops represents a prezygotic mating barrier that prevents heterospecific matings.

Estimation of the Plastic Zone in Fatigue via Micro-Indentation.

Accurate knowledge of the plastic zone of fatigue cracks is a very direct and effective way to quantify the damage of components subjected to cyclic loads. In this work, we propose an ultra-fine experimental characterisation of the plastic zone based on Vickers micro-indentations. The methodology is applied to different compact tension (CT) specimens made of aluminium alloy 2024-T351 subjected to increasing stress intensity factors. The experimental work and sensitivity analysis showed that polishing the surface to #3 µm surface finish and applying a 25 g-force load for 15 s produced the best results in terms of resolution and quality of the data. The methodology allowed the size and shape of both the cyclic and the monotonic plastic zones to be visualised through 2D contour maps. Comparison with Westergaard's analytical model indicates that the methodology, in general, overestimates the plastic zone. Comparison with S355 low carbon steel suggests that the methodology works best for alloys exhibiting a high strain hardening ratio.

Determination of Mechanical and Fracture Properties of Silicon Single Crystal from Indentation Experiments and Finite Element Modelling.

It is well-known that cracks are observed around the impression during indentation of brittle materials. The cracks inception depends on load conditions, material and indenter geometry. The paper aims to use experimental micro-indentation data, FE simulations with cohesive zone modelling, and an optimisation procedure to determine the cohesive energy density of silicon single crystals. While previous studies available in the literature, which use cohesive zone finite element techniques for simulation of indentation cracks in brittle solids, tried to improve methods for the evaluation of material toughness from the indentation load, crack size, hardness, elastic constants, and indenter geometry, this study focuses on the evaluation of the cohesive energy density 2Γ from which the material toughness can be easily determined using the well-known Griffith-Irwin formula. There is no need to control the premise of the linear fracture mechanics that the cohesive zone is much shorter than the crack length. Hence, the developed approach is suitable also for short cracks for which the linear fracture mechanics premise is violated.

Influence of Cross-Linking Agent Concentration/Beta Radiation Surface Modification on the Micro-Mechanical Properties of Polyamide 6.

This study focuses on the problematic of polyamide 6 containing various concentrations of cross-linking agent that was exposed to electron radiation. It is important to improve the material properties of polymers as much as possible. This endeavor can be realized by numerous methods, one of which is radiation exposure. This study investigates the effect of electron beam radiation in doses ranging from 66 to 132 kGy on the micro-mechanical properties of polymers, specifically polyamide 6 filled with 1, 3 and 5 wt.% of cross-linking agent triallyl isocyanurate (TAIC). The changes in the material brought by the radiation exposure were quantified by measurements of indentation hardness and modulus, which were the main measured micro-mechanical properties. Furthermore, thermo-mechanical analysis (TMA) was chosen to confirm the results of the material cross-linking, while the effect of degradation was investigated by Fourier-transform infrared spectroscopy (FTIR). In pursuit of complete evaluation, the topography of the test subject's surface was explored by atomic force microscopy (AFM). The optimal concentration/radiation ratio was found in polyamide 6 enriched by 5 wt.% concentration of TAIC, which was irradiated by 132 kGy. Material treated in such a way had its indentation hardness by 33% and indentation modulus improved by 26% in comparison with the untreated material. These results were subsequently confirmed by the TMA and FTIR methods.

The measured mechanical properties of osteoporotic trabecular bone decline with the increment of deformation volume during micro-indentation.

PURPOSE: Osteoporosis is a critical global health issue. However, the biomechanical properties of osteoporotic trabecular bone have not been well understood due to its hierarchically complex structure mingled with accumulated microcracks. Previous studies indicated the mechanical behaviors of trabecular bone may differ with varying amounts of deformation. Therefore, this study aims to further reveal the relationship between the measured mechanical properties of osteoporotic trabecular bone and various amounts of deformation volume during micro-indentation. METHODS: Two trabecular specimens were dissected transversally and frontally from an osteoporotic lumbar vertebral (L5) cadaver and embedded into Methyl methacrylate. On each specimen, two orthogonal cuts were performed to make a right-angle, followed by five parallel slicing. On each slice, the region of interest was gridded into 16 (4 × 4) sub-regions with the size equal to the microscope field. Within each sub-region, indentations were made on a single trabecula with five different indentation depths (3, 4, 5, 6, 7 µm) to induce different deformation volume. Both the indentation hardness and modulus were computed from the indenting curve for each measurement. The results of the five slices are pooled together to represent the longitudinal and circumferential mechanical characteristics, respectively. Linear regression was performed to investigate the relationship between the measured mechanical properties and various deformation volumes. RESULTS: A total of 1055 indents were made. After eliminating outliers, 840 indents were left for data analysis with 490 indents from transversal slices and 350 indents from frontal slices. Both the hardness and modulus decreased with the increasement of indentation depths. The hardness decreased by slopes of -0.65 (R2 = 0.72, p = 0.044) and -0.869 (R2 = 0.95, p = 0.003) longitudinally and circumferentially while the modulus decreased by slopes of -0.39 (R2 = 0.82, p = 0.02) and -0.348 (R2 = 0.94, p = 0.004) longitudinally and circumferentially. CONCLUSIONS: Mechanical properties of trabecular bone measured by micro-indentation can alter with the variation of deformation volume, which reflects the nonlinear behavior of vertebra from the material perspective.

Applications of Micro-Indentation Technology to Estimate Fracture Toughness of Shale.

The fracture toughness of shale is a basic parameter that can provide effective theoretical support for wellbore stability and hydraulic fracturing of a shale reservoir. Due to the composition and microstructure, there are many problems in evaluating the mechanical properties of shale in a macroscopic test. In this paper, the composition and pore distribution of shale were studied by X-ray diffraction and nuclear magnetic resonance. Scanning electron microscopy was used to characterize the pore structure. The setting of experimental parameters and the selection of the indenter were discussed. Micro-indentation technique was proposed and applied to fracture toughness analysis of shale. The results show that Berkovich indenter is more suitable for shale indentation test than Vickers indenter. Fracture toughness of shale indentation is obviously affected by surface roughness and indentation position. Fracture toughness of shale decreases slightly with the increase of the indentation load. The energy analysis result presents that the effect of cracking on the ratio of total/unloading work is minimal when there is no significant stripping on the shale surface. Compared with the experimental method, energy methods can obtain all the analysis parameters from a single indentation test. The results of comparative analysis with macroscopic experiments display that micro-indentation test can effectively predict the macroscopic fracture toughness of shale.