Near infrared spectroscopic assessment of loosely and tightly bound cortical bone water.

Water is an important component of bone and plays a key role in its mechanical and structural integrity. Water molecules in bone are present in different locations, including loosely or tightly bound to the matrix and/or mineral (biological apatite) phases. Identification of water location and interactions with matrix components impact bone function but have been challenging to assess. Here, we used near infrared (NIR) spectroscopy to identify loosely and tightly bound water present in cortical bone. In hydrated samples, NIR spectra have two primary water absorption bands at frequencies of ∼5200 and 7000 cm-1. Using lyophilization and hydrogen-deuterium exchange assays, we showed that these absorption bands are primarily associated with loosely bound bone water. Using further demineralization assays, thermal denaturation, and comparison to standards, we found that these absorption bands have underlying components associated with water molecules tightly bound to bone. In dehydrated samples, the peak at ∼5200 cm-1 was assigned to a combination of water tightly bound to collagen and to mineral, whereas the peak at 7000 cm-1 was exclusively associated with tightly bound mineral water. We also found significant positive correlations between the NIR mineral absorption bands and the mineral content as determined by an established mid infrared spectroscopic parameter, phosphate/amide I. Moreover, the NIR water data showed correlation trends with tissue mineral density (TMD) in cortical bone tissues. These observations reveal the ability of NIR spectroscopy to non-destructively identify loosely and tightly bound water in bone, which could have further applications in biomineralization and biomedical studies.

Environmentally-Controlled Near Infrared Spectroscopic Imaging of Bone Water.

We have designed an environmentally-controlled chamber for near infrared spectroscopic imaging (NIRSI) to monitor changes in cortical bone water content, an emerging biomarker related to bone quality assessment. The chamber is required to ensure repeatable spectroscopic measurements of tissues without the influence of atmospheric moisture. A calibration curve to predict gravimetric water content from human cadaveric cortical bone was created using NIRSI data obtained at six different lyophilization time points. Partial least squares (PLS) models successfully predicted bone water content that ranged from 0-10% (R = 0.96, p < 0.05, root mean square error of prediction (RMSEP) = 7.39%), as well as in the physiologic range of 4-10% of wet tissue weight (R = 0.87, p < 0.05, RMSEP = 14.5%). Similar results were obtained with univariate and bivariate regression models for prediction of water in the 0-10% range. Further, we identified two new NIR bone absorbances, at 6560 cm-1 and 6688 cm-1, associated with water and collagen respectively. Such data will be useful in pre-clinical studies that investigate changes in bone quality with disease, aging and with therapeutic use.

Validated Approaches for Quantification of Bone Mineral Crystallinity Using Transmission Fourier Transform Infrared (FT-IR), Attenuated Total Reflection (ATR) FT-IR, and Raman Spectroscopy.

Bone mineral crystallinity is an important factor determining bone quality and strength. The gold standard method to quantify crystallinity is X-ray diffraction (XRD), but vibrational spectroscopic methods present powerful alternatives to evaluate a greater variety of sample types. We describe original approaches by which transmission Fourier transform infrared (FT-IR), attenuated total reflection (ATR) FT-IR, and Raman spectroscopy can be confidently used to quantify bone mineral crystallinity. We analyzed a range of biological and synthetic apatite nanocrystals (10-25 nm) and found strong correlations between different spectral factors and the XRD determination of crystallinity. We highlight striking differences between FT-IR spectra obtained by transmission and ATR. In particular, we show for the first time the absence of the 1030 cm-1 crystalline apatite peak in ATR FT-IR spectra, which excludes its use for analyzing crystallinity using the traditional 1030/1020 cm-1 ratio. The ν4PO4 splitting ratio was also not adequate to evaluate crystallinity using ATR FT-IR. However, we established original approaches by which ATR FT-IR can be used to determine apatite crystallinity, such as the 1095/1115 and 960/1115 cm-1 peak ratios in the second derivative spectra. Moreover, we found a simple unified approach that can be applied for all three vibrational spectroscopy modalities: evaluation of the ν1PO4 peak position. Our results allow the recommendation of the most reliable analytical methods to estimate bone mineral crystallinity by vibrational spectroscopy, which can be readily implemented in many biomineralization, archeological and orthopedic studies. In particular, we present a step forward in advancing the use of the increasingly utilized ATR FT-IR modality for mineral research.

Lung Cancer: Are we up to the Challenge?

Lung cancer is the leading cause of cancer deaths worldwide among both men and women, with more than 1 million deaths annually. Non-small cell lung cancer (NSCLC) accounts for about 80% of all lung cancers.Although recent advances have been made in diagnosis and treatment strategies, the prognosis of NSCLC patients is poor and it is basically due to a lack of early diagnostic tools.However, in the last years genetic and biochemical studies have provided more information about the protein and gene's mutations involved in lung tumors. Additionally, recent proteomic and microRNA's approaches have been introduced to help biomarker discovery.Here we would like to discuss the most recent discoveries in lung cancer pathways, focusing on the genetic and epigenetic factors that play a crucial role in malignant cell proliferation, and how they could be helpful in diagnosis and targeted therapy.