| Intracerebral hemorrhage?ICH?is associated with high mortality and disability,which has attracted widespread attention from both domestic and overseas scholars.The onset and development of ICH are affected by the mass effect,the coagulation cascade,hemoglobin breakdown products,the related inflammation and genetic factors.Currently,the basic and clinical researches both focus on the mechanisms and therapy approaches of neural injuries induced by the coagulation cascade,hemoglobin breakdown products and the inflammation after ICH.But the mass effect also plays an important role in ICH.It changes the brain's architecture and even leads to life-threatening brain herniation,and has a close relationship with high mortality and poor outcome.However,the effect of mass effect on brain injuries and the related pathways were not clear.Therefore,exploring the effect of mass effect after ICH could disclose the development of ICH and provide new strategies for intervention.In the present study,the in vitro controllable hydrostatic pressure-loaded cell model and in vivo mass effect-induced rat ICH model were established based on the hydrostatic pressure induced by mass effect from hematoma and edema after ICH using the biomechanical strategies.The effect of hydrostatic pressure on cells and neural tissues,mechanotransduction protein,and cooperative action of hydrostatic pressure plus hemoglobin were detected.The results could supply a basic understanding for the influence of hydrostatic pressure derived from mass effect on perihematomal tissues after ICH.Furthermore,the function of mechanotransduction protein in vitro was investigated by the addition of inhibitor in the cell culture medium,and nicardipine hydrochloride?NCD?was delivered to brain tissues of ICH rat by nasal delivery system to investigate the inhibitory effect of mass effect on the perihematomal tissues via mechanotransduction protein inhibition,which could provide new strategies for ICH intervention.The main contents and results of present study are as follows:The in vitro controllable hydrostatic pressure-loaded cell model was constructed to explore the effect of hydrostatic pressure on the primary cortical neurons.Meanwhile,based on the volume of hematoma and edema after ICH in clinical practice and our preliminary finite element analysis,we chose kPa level of hydrostatic pressure loading on neurons.The neurons injuries were more seriously with the increase of hydrostatic pressure?20,30 and 40 kPa?.Compared to the control group,the decreased neurons viability,increased LDH releasing,damaged neuron architecture and increased cell apoptosis were observed when exposing to the high level of hydrostatic pressure?40kPa??*p<0.05?.Meanwhile,the cooperative effects of hydrostatic pressure?40 kPa?and hemoglobin had a significant difference compared with the exclusive hydrostatic pressure group?*p<0.05?.Results indicated that the elevated hydrostatic pressure induced by mass effect could insult neuron viability and apoptosis significantly after ICH.Furthermore,different volumes of agarose gel were injected into the right striatum of SD rats to simulate the mass effect on the perihematomal tissues after ICH in vivo and investigate the cooperative effects of mass effect and hemoglobin.Firstly,by MRI and images of coronal plane,we found that the mass volume was associated with the gel volume?25,50 and 100?L?,and the induced hemorrhage was significantly lower than the autologous whole blood-injection ICH model?**p<0.01?.Therefore,the mass effect-induced rat ICH model in this study could be used to research the effect of hydrostatic pressure induced by the exclusive mass effect on neural tissues,and it largely avoid the influence of hemoglobin.Secondly,results indicated that the hydrostatic pressure could disrupt the architecture of brain tissues,decrease the neural activity,increase brain edema and neural apoptosis surrounding the mass effect of agarose gel in the in vivo model.Lastly,the larger volume of gel insulted the brain tissues much more seriously.The hydrostatic pressure also exacerbated the neural injuries from hemoglobin.Therefore,results indicated that mass effect from the hematoma and brain edema not only gave rise to brain injuries directly in the acute stage of ICH,but also contributed to the following injuries induced by hemoglobin in the secondary brain injury after ICH.Besides,the larger volume of mass effect induced more serious brain injuries.The above results supply the theoretical references for the effect of hydrostatic pressure on the perihematomal tissues which induced by the mass effect from hematoma and edema after ICH.By the in vitro and vivo models,we explored a critical protein which participated in the neural injuries induced by hydrostatic pressure.Firstly,the protein expression of Piezo-2 was increased by the exclusive hydrostatic pressure?40 kPa?significantly in vitro?*p<0.05?.Though the exclusive hemoglobin had no significant effect on Piezo-2expression,its cooperative effect with hydrostatic pressure induced a significantly higher Piezo-2 expression compared with the exclusive hydrostatic pressure?*p<0.05?.So the hydrostatic pressure exhibited a synergistic effect with hemoglobin.Secondly,the mass effect increased the expression of Piezo-2 in a volume-dependent manner and the cooperative effect with hemoglobin increased its expression further in vivo.Lastly,both of nicardipine hydrochloride?NCD?and GsMTx-4 could decrease the protein expression of Piezo-2 which was increased by the hydrostatic pressure and inhibit Ca2+channel in vitro.They also reduced the neuron injuries from hydrostatic pressure which was mediated by Piezo-2,increased neurons viability and decreased neurons apoptosis significantly?*p<0.05?.The above results showed the related pathways of neural injuries induced by hydrostatic pressure after ICH.To improve the neural injuries in the acute stage after ICH,and based on the rapid drug absorption by intranasal delivery system,a nanoparticles-loaded in situ gel delivery system was developed in the present study to inhibit the expression of Piezo-2and decrease the neural injuries induced by hydrostatic pressure.Meanwhile,the in situ gel was coated with different charged surfaces to facilitate the release of charged nanoparticles from the in situ gel by electrostatic interactions.Firstly,the charge-driven transport of nanoparticles in gel with positively charged surfaces was performed in vitro.Results indicated that about 90%of positively charged nanoparticles could accumulate to the top of gel?the opposite side of the charged surfaces?.Meanwhile,the in vitro drug release studies and in vivo pharmacokinetic studies indicated that the charged surfaces could facilitate the transport of nanoparticles in gel and the release of drug to brain tissues.Secondly,results indicated that the in situ gel could keep nanoparticles on the nasal mucosa for extended periods of time,and the charged surfaces enhanced the absorption of drug and improved drug bioavailability?increased by 70%?by the electrostatic interactions.Thirdly,the released NCD could suppress the expression of Piezo-2 and inhibit the effect of hydrostatic pressure mediated by Piezo-2.Lastly,NCD could reduce the volume of mass effect from hematoma and brain edema,improve the neural injuries induced by hydrostatic pressure,maintain the brain's architecture and decrease neural apoptosis.The above experiments indicated that NCD could decrease the neural injuries derived from hydrostatic pressure by inhibiting the expression of Piezo-2.Therefore,exploring the effect of mass effect in ICH development could clarify the development of ICH and provide new strategies for clinical intervention. |
PDF Full Text Request